Congress 2019

Eugene CHRISTO-FOROUX: Mollivirus Kamchatka : a second representative of the mollivirus familly provides better insights into molliviridae evolution

In a context of global warming, deep investigations of viruses retaining their infectivity in prehistorical permafrost layers led to the discovery in 2015 of a 30,000-y-old fourth type of giant virus, Mollivirus sibericum. The assessment of the infectious threat contained in Russian soil is still an ongoing project. The analyses of modern samples from the Kamchatka region using both metagenomic approach and giant virus reactivation experiments allowed us to isolate a new virus called Mollivirus Kamchatka. These two combined approaches permit a reliable appraisal of the residual infectious threat of both bacterias and DNA viruses contained in the melting permafrost.


We can already confirm that the Mollivirus familly has not gone extinct and has expanded at least accross the Russian Federation. Those two representatives of these Acanthamoeba-infecting large DNA viruses family are showing a similar nucleo-cytoplasmic replication cycle as previously described for Mollivirus sibericum. Virions display a characteristic spherical shape ∼0.6 μm in diameter respectively enclosing a 648-kb to 651-kb fully syntenic GC-rich genome. Genetic features display 495 predicted proteins for Mollivirus sibericum and 480 proteins for Mollivirus Kamchtka of wich an average of 62 % are familly-specific ORFans.


Strain specific genes shows recent horizontal gene transfers between Mollivirus and Pandoravirus family. Results suggest that most of the genome is under negative/purifying selection and must have thus significantly contributed to virus' fitness over the last 30,000 past years. As a conclusion we discuss the de novo gene creation process observed in Pandoravirdae1 and the relevance of current viral classification.

Ao ZHANG: Functional characterization of the Bacillus subtilis PrkA protein

The enzymes that transfer a phosphate group from a molecule of ATP onto a protein are known as protein kinases. Protein phosphorylation is important for the regulation of a wide range of cellular processes both in eukaryotes and in prokaryotes (1). In bacteria, kinases shown to catalyze only phosphorylation of serine and threonine residues were thus referred as Ser/Thr Protein Kinases (STPKs) (2). There are four of these STPKs in Bacillus subtilis named PrkA, PrkC, PrkD and YabT. During the process of sporulation, they are reported to have important roles (3). Among these kinases, PrkA has been proved to significantly affect the rate of sporulation. However, the mechanism of this effect has not been revealed yet.


The biochemical properties of PrkA are uncharacterized but this protein possesses 631 amino-acids forming two domains: a N-terminal ATPase domain with Walker A and B motifs and a C-terminal STPK domain (Kinase domain). We cloned the full-length gene and truncated versions of prkA corresponding to the separated domains of PrkA as well as mutated versions potentially affected in one or the other activity of PrkA protein. We transferred these constructions into the prkA deletion strain to construct new strains for which the sporulation rate was calculated. The results showed that the deletion of prkA decreased about 20% the rate of sporulation. Complementation experiments with the separated domains showed that only the Kinase domain was able to rescue the ability to produce spores with a rate similar to the WT strain. Important residues for the kinase activity were also revealed by this approach. We then purified the full-length protein as well as the potential ATPase and Kinase domains by affinity purification procedure. The biochemical properties of the several proteins are now tested but further work must be done to understand the role of PrkA in the sporulation process in B. subtilis.

Nian Liu: Cellobiose phosphorylase CbpA is essential for cellobiose and cellulose catabolism in Ruminiclostridum cellulolyticum

Cellulose is the most abundant polysaccharide produced on earth. This polymer of glucose represents a large reservoir of glucose and a remarkable potential renewable source of energy. However glucose units are hardly accessible from cellulose due to its crystalline structure. Still, some bacteria are able to grow on this material, like the model Ruminiclostridium cellulolyticum. This bacterium is able to degrade cellulose into cellodextrins of various sizes (G5 to G2) and to import them in the cytosol where they are subsequently degraded by unknown process. We expected that phosphorolysis might be the preferred route of intracellular degradation of the cellodextrins over hydrolysis. Indeed it is energetically advantageous since this process releases phosphorylated glucose from cellodextrins that can directly enter the glycolysis pathway without need of extra ATP molecule for its phosphorylation as compared to hydrolysis pathway.

In this study, we addressed the role of 4 identified phosphorylases, CbpA, CdpA, CdpB and CdpC in the intracellular steps of degradation of cellulose, studying them at biochemical and genetic level. We showed that all the enzymes tested were highly active excepted CdpB. CdpA is specific to G5 and G4, CdpC is specific to G3, and CbpA is a strict cellobiose (G2) phosphorylase releasing glucose and glucose-1-phosphate. Combined, these enzymes are able to sequentially degrade G5 in 1 glucose and 4 glucose-1-phosphate. Based on gene inactivation of the phosphorylases genes and complementation studies, CbpA was found to be essential for cellobiose and cellulose catabolism in R. cellulolyticum.

Giuseppina R. BRIOLA : Studying dynamics and function relationship in HCPR: innovative approaches by incorporation of ncAAs, SDSL and EPR spectroscopy

Cellulose is the most abundant polysaccharide produced on earth. This polymer of glucose represents a large reservoir of glucose and a remarkable potential renewable source of energy. However glucose units are hardly accessible from cellulose due to its crystalline structure. Still, some bacteria are able to grow on this material, like the model Ruminiclostridium cellulolyticum. This bacterium is able to degrade cellulose into cellodextrins of various sizes (G5 to G2) and to import them in the cytosol where they are subsequently degraded by unknown process. We expected that phosphorolysis might be the preferred route of intracellular degradation of the cellodextrins over hydrolysis. Indeed it is energetically advantageous since this process releases phosphorylated glucose from cellodextrins that can directly enter the glycolysis pathway without need of extra ATP molecule for its phosphorylation as compared to hydrolysis pathway.

In this study, we addressed the role of 4 identified phosphorylases, CbpA, CdpA, CdpB and CdpC in the intracellular steps of degradation of cellulose, studying them at biochemical and genetic level. We showed that all the enzymes tested were highly active excepted CdpB. CdpA is specific to G5 and G4, CdpC is specific to G3, and CbpA is a strict cellobiose (G2) phosphorylase releasing glucose and glucose-1-phosphate. Combined, these enzymes are able to sequentially degrade G5 in 1 glucose and 4 glucose-1-phosphate. Based on gene inactivation of the phosphorylases genes and complementation studies, CbpA was found to be essential for cellobiose and cellulose catabolism in R. cellulolyticum.

Hui SHAO : Exploring the physiological regulation and crystal structure of CP12s in Thalassiosira pseudonana

Diatoms are photosynthetic organisms found in various aquatic environment and also in charge of one-fifth of the primary productivity on earth (Field et al, 1998). Carbon fixation plays key role in photosynthesis, which was regulated by a small protein, CP12, and the complex it formed with another two enzymes in green algae. CP12 is an intrinsically disordered protein (IDP) of 8,2-8,5 kDa and involved in many functions (Gontero and Maberly, 2012). Whereas the regulation of CP12 in diatoms remains unclear.

Thalassiosira pseudonana was the first species to be performed whole genome sequencing in diatoms (Armbrust et al, 2004). Previous study admitted that the expression of CP12 increased under low concentration of CO2 in T. pseudonana (Clement et al, 2017). My work consists in its physiological function and crystal structure. To acknowledge its characteristics, we investigated the recombinant wild-type protein by nuclear magnetic resonance, gel filtration, small angle X-ray scattering and circular dichroism. Results showed it’s perhaps a spherical oligomer. Heat treatment displayed it belongs to IDPs. Few observed changes of reduced form by dithiothreitol may suggest the other regulation instead of by thioredoxin. Western blot and mass spectrometry showed the expression under dark condition in vivo. Further studies need to be addressed to the regulation with other enzymes and more expressed environment in nature.

Hanna BISMUTH : Galleria mellonella: Characterization of the oxidative stress response during infection by Salmonella enterica

Bacteriophages (phages) are viruses that specifically infect bacteria. Some of them can integrate their DNA into the bacterial chromosome; they are then called prophages. Most prophage genes are silent, but those that are expressed can provide new properties to their host.

In E. coli K12, the AppY protein encoded by the DLP12 prophage has a dual role in the cell: it’s a transcriptional regulator from the AraC family as well as a new factor able to stabilize RpoS. RpoS is the major sigma factor during stationary phase and under several stress conditions. Its accumulation in the cell leads to the transcription of more than 500 genes. When not needed, RpoS interacts directly with the adaptator protein RssB, which brings it to degradation by the ClpXP protease. It has been shown in the laboratory that AppY can block this degradation pathway by titrating RssB, leading to RpoS stabilization in the cell.

To understand the role of AppY as a transcriptional regulator in the bacterial physiology, a RNA-Seq experiment has been performed. This technique has allowed us to identify new AppY’s targets involved in acid stress resistance and biofilm formation. We are currently characterizing further AppY involvement in these two pathways. Moreover, by performing a genetic screen we have already demonstrated that the two functions of AppY can be separated by identifying specific residues involved in each of them.

The goal of this work is to understand how AppY influences the E. coli physiology and how prophages co-evolve with their host.

Mohamed MROUEH / Sandrine PAGES : The arabinoxylan metabolism in Ruminiclostridium cellulolyticum

The α-L-Arabinofuranosidases (α-L-ABFs) are exo-enzymes involved in the hydrolysis of -L-arabinosyl linkages in plant cell wall polysaccharides. They play a crucial role in the degradation of arabinoxylan, a substrate highly decorated by arabinose residues and acetyl groups. R. cellulolyticum produces four cellulosomal -L-ABFs belonging to families 43 and 62 of the glycoside hydrolases (GH), encoded by the xyl-doc gene cluster (loci Ccel_1229, Ccel_1233, Ccel_1234 and Ccel_1240). GH43-1229 presents a low specific activity on natural substrates and could cleave off arabinose decorations located at arabinoxylan chain extremities. GH43-1233 is able to convert the double arabinose decorations into single O2- or O3-linked decorations with high velocity (kcat = 16.6 ± 0.59 s-1) and acts in synergy with GH62-1234 which hydrolyzes -arabinosyl linkages on monodecorated substrates. Finally a bifunctional enzyme, GH62-CE6-1240, containing a GH62--L-ABF module and a Carbohydrate Esterase (CE6) module, catalyzes the deacylation of substrate and the cleavage of -arabinosyl monosubstitutions. All together these enzymes are able to remove the arabinoxylan decorations allowing the following action of endoxylanases. Products of the arabinoxylan degradation, arabinose and xylose might be imported by the clostridia by the way of an ABC-transporter specific for monosaccharides, encoded by genes upstream of the xyl-doc cluster (loci Ccel_1223 to Ccel_1226). This sophisticated extracellular degradation system might even be completed by a second system encoded by a 13-gene cluster now under study, which is predicted to directly manage small arabinoxylodextrins import and degradation in the cytosol, thus increasing further the complexity of arabinoxylan catabolism.

Asmaa HADJ AHMED : The design of a new electrochemical cell for studying enzymes that reduce CO2

In our Team in BIP Laboratory, Direct electrochemistry (DE) is utilized to study a number of metalloenzymes which catalyze redox reactions of interest, such as the interconversion between protons and H2 or the reduction of CO2 to CO. In this technique, the enzyme is immobilized on a rotating disc electrode in a configuration where the electron transfer is direct, and the enzymatic activity is monitored as an electrical current. This technique has proved extremely useful to study various aspects of the reactivity of metalloenzymes [1]. However, a problem appears when trying to use it to study CO dehydrogenase (CODH), which catalyses the reversible reduction of CO2 to CO following this equation:

CO2 + 2e-+ 2H+ <=> CO + H2O

This enzyme is so fast that the catalysis is mostly limited by the transport of the substrate (CO) towards the electrode, and not by the chemical reaction, which blurs all the mechanistic information in the electrochemical signals [2]. This problem couldn't be solved using the rotating disc electrode even at the maximal rotation rate. Hence, we need to change the setup and design a new electrochemical cell with improved transport properties. We develop an approach in which we use computational fluid dynamics to screen for suitable geometries with improved transport properties, which we will then build and use for mechanistic. In this work, we will present the different cell geometries that we have screened and their transport properties deduced from numerical simulations.

Ivy MALLICK : APEX2 as a potential genetic tag for targets identification involved in pathogenic mycobacterial persistence

With 10.4 million new cases and 1.7 million deaths worldwide, tuberculosis (TB) has been considered as the deadly infectious disease alongside with HIV. Establishment of the dormancy state and reactivation of the disease are critical to the infectious-lifecycle of Mycobacterium tuberculosis (Mtb). During infection, in granulomas, among the different cells, one important subpopulation is the foamy macrophages (FM), characterized by the abundance of triacylglycerol-rich lipid bodies (LB). In these FM, Mtb can reside, where it acquires free lipids from host’s LB, subsequently processed and stored by the bacilli in the form of intra cytosolic lipid inclusions (ILI) leading to dormancy. The molecular mechanisms and the actors whereby intra phagosomal mycobacteria interact with LB, can assimilate the LB-derived lipids to form ILI and hydrolyze TAG from ILI during the reactivation are poorly understood.

Live cell tagging by different agents has major limitations like long incubation time or long half-life or inactivation inside the live cell environment. One solution to circumvent these limitations is the use of APEX2 as compartment specific live cell genetic tag, which is getting popular now-a-days. APEX2 is an engineered variant of the monomeric ascorbate peroxidase that oxidizes biotin-phenol to short-lived (<1 msec), small-distance (<20 nm) diffusive biotin-phenoxyl radicals in the presence of hydrogen peroxide, that covalently reacts with amino-acid side chains of neighboring proteins allowing biotinylation. These proximity-biotinylated proteins can then be enriched on streptavidin and identified by mass spectrometry.

So the specific aim of the study includes deciphering the mechanism of utilization of host’s lipids by Mycobacterium to accumulate TAG in the form of ILI inside foamy macrophages and determining how the bacteria is able to consume these lipids during the reactivation. We expect that identification of proteins (host and mycobacterial) involved either in the formation and degradation of ILI in in vitro or ex vivo situations will enlighten our knowledge about the reactivation, persistence and dormancy processes.

In our approach, three proteins (Tgs1, Rv1683 and HBHA) already known to interact with the ILI will be fused to APEX2 and only protein targets in the vicinity of the Tgs1-APEX2, Rv1683-APEX2 or HBHA-APEX2 fusions will be streptavidin/anti-biotin purified, based on the presence of biotinylated chemical functions. Therefore, Applying such new approach and methodology will allow us to decipher the physiological role of these ILIs and identify new enzymes or protein partners involved in ILI formation/degradation, providing new insights into mycobacterial persistence and latent TB infection.

Siyi LIU : Iron-sulfur biogenesis

Iron-sulfur (Fe-S) clusters are ancient cofactors that exist in all kingdoms of life. They are involved in proteins functioning in a wide range of physiological processes including gene expression, DNA repairation, central metabolism and respiration. Fe-S clusters are essential for all living organisms. ISC machinery is responsible for Fe-S biogenesis in both prokaryotes and eukaryotes.

Frataxin (FXN) is part of the Fe-S assembly complex. The role of FXN is debated. Previous studies have suggested it as an iron donor, a regulator in the process of cluster assembly or a member of reactive oxygen species (ROS) detoxification. Deficiency of FXN results in a serious neurodegenerative disease Friedreich’s ataxia in human beings and strong phenotype in yeast. How to by-pass FXN? To answer this question, we searched for suppressors.

Anne BOYELDIEU : An unexpected regulatory network including several diguanylate cyclases controls biofilm formation in Shewanella oneidensis

Shewanella oneidensis is an aquatic proteobacterium capable of forming biofilms either associated to surfaces (SSA-biofilm) or at the air liquid interface (pellicle). We have previously shown that pellicle biogenesis in S. oneidensis requires the chemotaxis histidine kinase CheA3 and its cognate response regulator CheY31.


We recently identified two genes encoding putative diguanylate cyclases (DGCs) (pdgA and pdgB) that, when overexpressed, allow pellicle formation in the non‐pellicle‐forming cheY3‐deleted mutant2. A mutant deleted of both pdgA and pdgB is affected during pellicle development. This result indicated the involvement of the c-di-GMP secondary messenger in pellicle formation. Interestingly, we showed that one target of c-di-GMP is the MxdA protein, which interacts with both PdgA and CheY3. Since the mxdA gene belongs to an operon previously proposed to be involved in exopolyssacharide biosynthesis, we propose that this complex regulatory network promotes pellicle biogenesis by allowing the biosynthesis of the matrix.


I have recently discovered that a cheY3-deleted strain is also impaired for SSA-biofilm formation. Indeed, a strain deleted of cheY3 is unable to adhere to solid surfaces. By using a multicopy library of S. oneidensis chromosomic fragments, I identified three novel diguanylate cyclases that allow pellicle and/or SSA-biofilm formation in the cheY3-deleted mutant. I am currently investigating the role of these DGCs in pellicle and SSA-biofilm development. I propose that these DGCs are temporally activated in response to specific signals during the different steps of biofilm biogenesis.

Bouchra ATTIA : Structural characterization of the GtlJ protein and its interactions with the cytoplasmic platform during the adventurous motility of Myxococcus xanthus

The motility of bacterial cells promotes a range of physiological phenomena such as nutrient detection, predation, biofilm formation and pathogenesis. Myxococcus xanthus uses the so-called adventurous motility (A-motility) to explore new places, change direction and leave the cell swarms.

All the studies available revealed that this motility is controlled by cytoplasmic proteins1–3, MglA and AglZ, coupled to the bacterial cytoskeleton mreb and a molecular motor agl, as well as a multiprotein complex4 (Glt) crossing the entire bacterial membrane. This assembly forms focal adhesion complexes (FACs) which propel the cell in a certain direction beforehand defined by the cytoplasmic protein, MglA. Within the Glt multiprotein complex, GltJ is a protein who plays a critical role in FACs assembly and regulation. GltJ is located in the inner membrane with a cytoplasmic n-terminus composed of a "zinc ribbon" (ZnR) domain and a "glycine-tyrosine-phenylalanine "(GYF) domain.

In order to understand at the molecular and structural level the underlying mechanism behind the A-motility, we used NMR spectroscopy to solve 3D structures of GltJ domains and decipher how the contacts between GtlJ domains and cytoplasmic proteins regulate the assembly of FACs. Our results not only represent the first high resolution structural data concerning the ZnR and GYF domains of the Glt complex but also highlight a direct contact between a cytoplasmic effector of the A-motility machinery and the complex Glt. These data allowed us to propose a role of molecular switch for the ZnR and GYF domains of GltJ regulating the assembly and disassembly of FACs.

Paul GUIRAUD : AriR activates the stringent response by triggering the alarmone synthetase SpoT in Escherichia coli.

The Stringent Response (SR) is a general bacterial stress response induced by diverse nutritional and environmental stresses allowing bacteria to adapt and survive adverse conditions. This reprogramming of cell physiology is caused by the accumulation of the alarmone (p)ppGpp which, in Escherichia coli, depends on two enzymes, the (p)ppGpp synthetase RelA and the bifunctional (p)ppGpp synthetase/hydrolase SpoT. While conditions that trigger SpoT-dependent (p)ppGpp accumulation have been described, the molecular mechanisms regulating the switch between the hydrolase and the synthetase activity remain poorly understood. In this work, we identified a new protein that activates stringent response in E. coli. By genetic screen assay we discovered that AriR (YmgB) promotes SpoT-dependent accumulation of (p)ppGpp in E. coli. Importantly, AriR controls the SR by direct binding to the TGS and Helical domains of SpoT, further supporting a pivotal role of these two domains in the reciprocal regulation of the two enzymatic states. AriR is a small protein which has been previously proposed to act as a connector for the phosphorelay Rcs involved in the biofilm formation and acidic stress response. Remarkably, by using random mutagenesis, we isolated amino acid substitutions in AriR that disrupt the AriR-SpoT interaction and abolish (p)ppGpp accumulation in vivo. Finally, one of those mutants (R74G) is still able to activate the Rcs phosphorelay showing that it is possible to dissociate the function of AriR on SpoT from its function on Rcs system.

Martino BENVENUTI : Electrochemical and biochemical comparison of two novel CODHs produced by the archaeon Thermococcus sp. AM4

Ni-Fe containing carbon monoxide dehydrogenases (CODH) are the biological catalysts that perform the reversible conversion of CO into CO2 according to the reaction

CO + H2O CO2 + 2H+ + 2e-

They are found in bacteria and archaea. Bacterial CODHs are the best characterized in terms of structure and catalytic properties while just few archaeal CODHs have been purified so far1,2. The active site of these enzyme, called the C-cluster, is a [Ni-4Fe-4S] center. The biosynthesis of this active site is not fully understood. In some cases, it depends on the accessory protein CooC3. Although Ni-Fe CODHs are highly similar in terms of structure, they have different catalytic properties. The affinity for CO and CO2 varies as well as the reactivity towards the inhibitor O24,5.

Here, we describe the productions and electrochemical and biochemical characterizations of two novel CODHs produced by the archaeon Thermococcus sp. AM4 (Tc-CODH I and Tc-CODH II). By purifying these CODHs, our aim is to understand the catalytic properties of archaeal Ni-Fe CODHs that are poorly characterized. Moreover, the direct comparison of these CODHs could help to understand their physiological role inside the cell. We heterogeneously produced and purified Tc-CODHs from Desulfovibrio fructosovorans in the presence or absence of the gene cooC that is present in the two CODH-encoding operons. When produced in the presence of CooC, the Ni-content of the two proteins increases and they can oxidize CO and reduce CO2. This result shows that they are CooC-dependent proteins. However, Tc-CODH II is much more active than Tc-CODH I in both catalyzing the oxidation of CO and reduction of CO2. We also show that Tc-CODH II is more sensitive than Tc-CODH I with the latter recovering part of its activity after reducing treatment.

Anna BONNARDEL : A new bacterial regulator controlling cell cycle and division in Sinorhizobium meliloti

Sinorhizobium meliloti is a gram-negative bacterium able to fix nitrogen in symbiosis with plants. It belongs to the class Alphaproteobacteria showing asymmetrical cell division at every cell cycle. In S. meliloti cell cycle and cell division are controlled by a complex regulatory network, in which, at the center, the master regulator CtrA plays a crucial role activating cell division, motility and repressing the replication of DNA. Among many regulators, the kinase DivJ acts as a repressor of CtrA activity. By a genetic screening, selecting for Tn mutations able to rescue the growth defect of delta-divJ, we identified an insertion in the C-terminal region of the gene we named DivJ growth suppressor (digS). We showed that complete deletion of digS is lethal in S. meliloti and cells depleted of DigS showed a dramatic “mini cells” phenotype, suggesting a clear cell division defect. We showed by His6-DigS affinity column that this new cell cycle factor is able to bind CtrA and FtsZ2. This link between DigS and CtrA and FtsZ2 was further supported by immunoblots against CtrA and FtsZ2, revealing that upon depletion of DigS the protein levels of the two interactors increase, suggesting a negative role of DigS on CtrA and FtsZ2. Finally DigS-GFP showed a clear mid-cell localization similar to the cell division septum consolidating the putative interaction with FtsZ apparatus.

In conclusion a new regulator plays a crucial role in coordination of cell cycle and cell division in S. meliloti and possibly in other members of the class Alphaproteobacteria, including Brucella and Agrobacterium.

Shuanghong XUE : Dif signalling pathway controls exopolysaccharide biosynthesis in Myxococcus xanthus

The bacterial exopolysaccharide (EPS) is the main component of extracellular matrix and it mediates fruiting body formation and type IV pilus dependent social motility in Myxococcus xanthus. The production of EPS is partially regulated by the Dif pathway, which is composed of homologs of the enteric chemotaxis system: a transmembrane receptor, DifA; a histidine kinase, DifE; a CheW-like, DifC and the core CheY, EpsW. The pathway includes an accessory CheY-like response regulator, DifD, which might function as a phosphate sink.

Our work shows that Dif might regulate, at post-transductional levels, the activity of the Wzx/Wzy-dependent polysaccharide assembly pathway (Wze) encoded by the Mxan7416-7421 gene cluster. The aim of my PhD project is to decipher how this Dif-mediates regulation of EPS production occurs. A first approach aims to analyze the cellular localization of selected Dif and Wze components to correlate localization patterns and functions.

A second approach consists of fishing the interacting partners of the Dif output, EpsW and DifD, by suppressor analysis and biochemistry by the aid of the newly developed APEX technique.

Christina FELBEK : Mechanistic studies of FeFe hydrogenases by protein film voltammetry

Our group seeks to understand the kinetics of hydrogenases during anaerobic and aerobic inactivation through protein film voltammetry (PFV) – a technique that has been developed and improved in the past to monitor enzyme activity. The time dependent change in activity (which is induced by e.g. a high potential) can be related to suitable kinetic models.[1] It has been shown that this technique leads to comparable kinetic information than the catalytic measurements of enzymes in solution.[2] Furthermore, with the help of DFT calculation the molecular structure of catalytic intermediates can be predicted. [3]

FeFe hydrogenases catalyze the proton reduction with an exceptional high turnover (100 molecules H2/s) making this group of enzymes interesting for the environmental friendly production of hydrogen fuel.[4] Using PFV, we seek to obtain a better understanding of these enzymes by comparing kinetics of of FeFe hydrogenases originating from different organisms and newly designed mutants. This project goes hand in hand with gaining further insight in the recently observed role of halides[5] and light[6] as inhibition factors for hydrogen oxidation.

Nicolas O. GOMEZ : Identification and characterization of a new zinc and nickel import pathway essential for Pseudomonas aeruginosa’s growth in lung mucus

Acquisition and fine-tuned homeostasis of divalent metals is a crucial process for every living organism, and particularly for bacteria in the framework of nutritional immunity encountered during infections. In order to highjack the metal containment strategy of the hosts, many pathogenic bacteria produce high affinity metal molecules called metallophores. We discovered, identified, and characterized a four-gene operon in Pseudomonas aeruginosa involved in the biosynthesis (cntL and cntM), inner membrane export (cntI) and outer membrane recovery (cntO) of a metallophore that we named pseudopaline (Lhospice et al. 2017). In vivo the pseudopaline operon is regulated by zinc through the repressor Zur. Functional studies revealed that pseudopaline is involved in nickel uptake in metal scarce media, and, most importantly, in zinc uptake in conditions mimicking a chelating environment. Considering that a functional pseudopaline pathway is essential for P. aeruginosa survival in airway mucus secretions (Gi et al. 2015), we are investigating which zinc-dependent protein, central to growth, pathogenicity and virulence in infection-like condition benefits from pseudopaline’s zinc intake for proper biogenesis and function. We observed a significant reduction of protease activity in the supernatant of a pseudopaline deficient strain. Further experiments revealed that the extracellular activity of the zinc-metalloprotease IMPa is severely impaired in metal scarce conditions, thus suggesting that this important virulence factor depend on pseudopaline during infection. Considering this pathway as a possible target for developing an antibacterial strategy, we synthetized two variants of pseudopaline presenting an inhibitory effect

Xiaomei XU : HetR Protection against PatS Inhibition, Mediated by HetL, is a Crucial Mechanism for Pattern Formation during Cell Differentiation in Anabaena PCC 7120

Anabeana PCC 7120, a filamentous and autotrophic cyanobacterium, is able to undergo cellular differentiation. Under sufficient supply of combined nitrogen, the whole filament is formed by vegetative cells; while with nitrogen deprivation, 5-10% of vegetative cells differentiate into distinctive heterocysts, responsible for nitrogen fixation and diazotrophic growth.


Multiple signals involved in regulatory mechanisms have been revealed. Among which, HetR, the master transcriptional regulator, initiates cell differentiation and activates several developmental genes. PatS, as the diffusible peptide, inhibits HetR in the vegetative cells, ensuring that only 5-10% of them enter the developmental program in a semiregular manner. HetR and PatS represent an example of lateral inhibition mechanism that explains the establishment of differentiation pattern.


With PatS being produced in heterocysts, one of the most important enigma to be solved is: what protects HetR from PatS inhibition in heterocysts? We have discovered that HetL, involved in the differentiation process, interacts with HetR without disrupting DNA-binding activity of HetR. Interestingly, HetL interacts with HetR at the same interface than PatS. Moreover, our data showed that PatS interferes with HetL-HetR interaction. A HetR variant which is insensitive to PatS, loses the interaction with HetL as well. In addition, while HetL overproduction in vivo abolished the inhibiting effect of PatS, a HetL variant unable to interact with HetR, was unable to sustain this protective effect. Altogether, it’s suggested that HetL ensures immunity to HetR against PatS in heterocysts. This result is an important step towards understanding how heterocyst pattern is made in Anabaena.

Dukas JURENAS : Molecular mechanism of regulation of ataR-ataT toxin antitoxin system

Toxin antitoxin (TA) systems are small genetic elements encoded in bacterial mobile genetic elements (MGE) and chromosomes. TAs are involved in stable inheritance mechanisms by inhibiting bacterial growth in the daughter cells that are losing the TA-bearing MGEs. Chromosomal TAs are thought to be involved in genetic programmes allowing to halt bacterial growth in unfavourable conditions. AtaT is a novel acetyltransferase toxin from AtaR-AtaT system from enterohemorrhagic Escherichia coli O157:H7. We have unraveled AtaT toxicity mechanism using in vivo and in vitro assays by tracking the transfer of acetyl moiety from Acetyl-CoA.

To study neutralization and transcription regulation we have obtained crystal structures of antitoxin, toxin, and a TA complex bound to DNA. We then confirmed specific interactions in vivo and in vitro by mutantions designed based on structural information. We have found AtaT blocks initiation of translation by acetylating initiator Met-tRNAfMet on methionine and thereby prevents its interaction with initiation factor IF2 and presentation to 30S ribosomal initiation complex. AtaT is neutralized by AtaR directly after its translation and locked in an inactive monomeric state. Heterohexameric AtaR2-AtaT-AtaR2-AtaT complex is an effective transcription repressor for ataR-ataT operon. Excess of AtaT breaks down the complex to heterotetrameric form. The later complexes can no longer repress transcription but still neutralize the toxin. AtaT when released form initial TA complex dimerizes and two monomers come together to form large positively charged binding surface that specifically recognizes its target tRNA.

Lisa ZUILY : Copper induces protein aggregation, role of a molecular chaperone

In the cellular environment, most proteins require a well-defined three-dimensional structure to fulfill their function. The process of protein folding is often guided bymolecular chaperones to avoid misfolding, aggregation, and the generation of toxic species. Here, we will focus on Hsp33, a redox-regulated molecular chaperone, highly conserved in bacteria. Hsp33 plays an important role in protecting cells against oxidative stress that lead to protein unfolding (1). Hsp33 is inactive under non-stress conditions, upon exposure to peroxide stress at elevated temperature (1) or to hypochlorous acid (2), Hsp33 gets rapidly activated. To respond to these stresses, Hsp33 possesses a sophisticated regulatory system with (i) a C-terminal redox switch domain which detects oxidizing conditions by disulfide bond formation and (ii) an adjacent metastable linker region which responds to unfolding conditions (3). Hsp33 prevents the aggregation of several client proteins and once normal condition resume, Hsp33 transfer its client to foldase chaperone (DnaK system) for refolding. Such stresses occur during infection, the immune system produces oxidative agent to kill invading bacteria. The release of toxic metal, like Cu, is also a strategy developed by the immune system to kill pathogenic bacteria. We are currently testing the impact of copper on E. coli cells and the role played by Hsp33 to protect proteins from metal toxicity. Little is known on the physiological role of molecular chaperones during metal stress and we hope to get more insight into this interesting topic.

Naoual DERDOURI : Involvement of AppY, a dual regulator from prophage origin, in the bacterial regulatory network

Bacteriophages (phages) are viruses that specifically infect bacteria. Some of them can integrate their DNA into the bacterial chromosome; they are then called prophages. Most prophage genes are silent, but those that are expressed can provide new properties to their host.

In E. coli K12, the AppY protein encoded by the DLP12 prophage has a dual role in the cell: it’s a transcriptional regulator from the AraC family as well as a new factor able to stabilize RpoS. RpoS is the major sigma factor during stationary phase and under several stress conditions. Its accumulation in the cell leads to the transcription of more than 500 genes. When not needed, RpoS interacts directly with the adaptator protein RssB, which brings it to degradation by the ClpXP protease. It has been shown in the laboratory that AppY can block this degradation pathway by titrating RssB, leading to RpoS stabilization in the cell.

To understand the role of AppY as a transcriptional regulator in the bacterial physiology, a RNA-Seq experiment has been performed. This technique has allowed us to identify new AppY’s targets involved in acid stress resistance and biofilm formation. We are currently characterizing further AppY involvement in these two pathways. Moreover, by performing a genetic screen we have already demonstrated that the two functions of AppY can be separated by identifying specific residues involved in each of them.

The goal of this work is to understand how AppY influences the E. coli physiology and how prophages co-evolve with their host.

Amaury PAYELLEVILLE : Whole genome DNA methylation (Methylome), transcriptomic and phenotypic analysis revealed involvement of Dam DNA methyltransferase in gene regulation in Photorhabdus luminescens.

DNA methylation is an epigenetic mechanism regulating genes expression by reducing the affinity of transcriptional regulators for their binding site (Casadesus and Low, 2006). Photorhabdus luminescens is an entomopathogenic bacteria switching from symbiosis, with a nematode, to pathogenicity, in the insect (Boemare et al., 1993). Because Dam methyltransferase, methylating GATC sites, is involved in pathogenicity of several bacteria, we decided to investigate its role in P. luminescens.

Methylome analysis revealed that 99% of GATC sites in the genome are methylated in all tested growth conditions. Overexpressing Dam methylates most of the unmethylated sites and causes a decrease in motility and pathogenicity of the bacteria whereas it increases biofilms formation. Transcriptomic analysis revealed that observed phenotypes are related to differences at transcriptional level. Coupling phenotypic, transcriptomic and methylomic analysis provides clues to identify genes transcriptionally regulated by DNA methylation and to understand Dam DNA methylation involvement in P. luminescens life-cycle.

Nicolas JOLIVET : Focal Adhesions in Gliding Bacteria are Stabilized By Accessory Adhesins in a Substratum-Dependent Manner

Contact-dependent communication in bacteria is a vital area of research, as bacteria often reside in communities in which detection of self-generated signals coordinate complex behaviours. This is of particular interest for bacteria that do not possess conventional quorum-sensing systems using autoinducer molecules. Gram-negative Myxococcus xanthus is used as model because it exhibits a “social” physiological lifecycle including group migration, biofilm and spore formation. One proposed method for coordinating these events is the contact-dependent recognition of polysaccharide deposed during single-cell motility (gliding). This motility involves transport of a gliding transducer complex (glt) from the leading to the lagging poles. Anchoring of the complex to the substratum involves an adhesin, which is proposed to interact with the polysaccharides. One other adhesin candidate is CglD, reported to be important for gliding.

Microscopy analyses of WT and ΔcglD cells revealed that CglD was dispensable for gliding on agar, but required for gliding on chitosan-coated glass slides in a Ca2+-dependent manner. Fold recognition analyses of CglD indicated a dual-domain structure; N-terminal match to the eukaryotic cartilage oligomeric matrix protein while the C-terminal match to the eukaryotic integrin. These proteins are widely implicated in adherence of eukaryotic cells. Microscopy of fluorescently-labelled Glt motility complexes in cells lacking CglD formed less stable bacterial clusters and also reversed their direction less frequently than WT cells. This may indicate an link between the stability of the gliding complex and the regulatory pathway responsible for cell polarity.

Alejandro VILLALTA : GG, an atypical iron-sulfur protein conserved in the Mimiviridae family

Giant viruses, since discovered 16 years ago, surprise by their unexpected features such as the size of the virions (>0.5µm), their complex genomes (600kb-2.5Mb), containing 650 to 2500 genes, most of them (>60%) encoding proteins without cellular or viral homologs. To date, four families have been identified: Mimiviridae, Pandoraviridae, Pithoviridae and Molliviridae (1) and the number of identified giant viruses is still increasing worldwide (2).


Transcriptional and proteomic analyses of Mimivirus, infecting Acanthamoeba castellanii, revealed that an unpredicted gene (R633b) was among the most transcribed at the late stage of the infection and encoded one of the most abundant proteins (GG) in the virions (3), showing its importance for the virus. GG is a small protein of ~6kDa conserved in the Mimiviridae, with an atypical sequence composed of ~40% glycine and 12% cysteine, coordinating an uncommon Fe-S cluster. Its function remains unknown.


Aiming to elucidate the structure and function of the GG-FeS family, we characterized the Fe-S cluster combining UV-Vis, EPR analyses of the wild type and mutant proteins. This unusual Fe-S cluster reminds the inactive aconitase’s linear 3Fe-4S (4) with the behavior of the IssA atypical thioferrate protein (5). Our structural studies show that GG can oligomerize to form 20nm structures that can assemble into long chains. The expression in Acanthamoeba castellanii of GG fused with GFP reveals its localization at the cellular membrane. I will also present some surprising properties of the protein that could serendipitously provide us the means to elucidate its 3D structure

Israel VERGARA ALVAREZ : Regulation and biosynthesis of exopolysaccharides in Myxococcus xanthus

Multicellularity plays a key role in bacteria lifespan. In fact, the formation of highly ordered structures confers benefits in terms of development and survival through communication and cooperation (Alegado & King, 2014). In the social δ-proteobacterium Myxococcus xanthus the formation of multicellular structures relies on the regulated biosynthesis of exopolysaccharides (EPS) by the Dif chemosensory system (CSS) (Yang et al., 2000) and three Wzx/Wzy dependent polysaccharide assembly machineries (Wz).

In this work, we identified, by bioinformatics, components of each machinery. We termed these machineries WzE for (Extracellular matrix); WzB for (Biosurfactant); and WzS for (Spore coat). We focus on WzE and WzB for which phenotypic analyses coupled to colorimetric assays suggest that while WzE is involved in the production of the main EPS important for Type IV pilus mediated motility and adhesion, WzB produces a sugar polymer with surfactant properties. To confirm our hypothesis, we isolated EPS from both Wz and carried out HPLC and measured their visco-elastic properties in a drop tensiometer. We show that Wz polymers syntethized have distinct composition and physical properties and, WzB titrates the adhesiveness of cells. Finally we design a trans-complementation experiment and showed that WT phenotype can be rescued, furthermore, by a spatio-temporal analysis of WzE and WzB promoters suggest that WzE is expressed at the cell border and the WzB at the colony center.

These data together reveal that the combined activity of two sugars allows community behaviors as observed in wild type cells.

Victoria SCHMIDT : The Pseudomonas aeruginosa T6SS membrane complex: from molecular dissection to chemical inhibition

Antibiotic resistance is a war for 21th century researchers. The WHO has classified some bacteria in critical priority for that reason and the development of new drugs to counteract them is crucial. Pseudomonas aeruginosa is one of these bacteria responsible for number of nosocomial infections; this ubiquitous opportunistic pathogens is a killer for immunodepressed patients and around 70% of Cystic Fibrosis patients succumb to this deadly bacteria.

One of the macromolecular systems involved in new ecological niches and hosts invasions is the type 6 secretion system (T6SS). Composed of 3 major sub-complexes (the membrane complex, the baseplate and the contractile tail), the T6SS act as a nano crossbow to inject toxic effectors in the target cell. The T6SS is well conserved structurally and found in various pathogens such as Acinetobacter baumannii, Vibrio cholerae, Klebsiella pneumoniae and P. aeruginosa.

To counteract the P. aeruginosa T6SS machinery we have to found a key interface between essentials components and potentially accessible from extracellular environment by small molecule inhibitors. One of the T6SS sub-complexes, the membrane complex (MC), seems to be an interesting target. The MC is the first part of the machinery being assembled, and is always present in the bacterial envelope, during and in between two T6SS firing cycles. Without this essential MC the rest of the machinery is not assembled. The outmost part of the MC is inserted into the outer membrane. The MC is an oligomer formed by 3 proteins : TssL, inserted in the cytoplasmic membrane, TssM, a huge protein inserted in both cytoplasmic and outer membranes, and TssJ a lipoprotein anchored in the outer membrane. The interface between TssJ and the periplasmic part of TssM could be an interesting target for T6SS inhibitors.

Our project is to design reporter tools to set-up a medium-to-high-throughput screening of chemical compounds that, by blocking the TssJ-TssM interface, could inhibit the P. aeruginosa T6SS. Using an in vitro HTRF screening on a specialized chemical library, our collaborators at Paoli Calmette institute (CRCM-IPC) have already identified several compounds that possibly target the TssJ-TssM interface (JM-PPI). To test these JM-PPI molecules we have developed various in vivo protein-protein interaction assays and, with the help of the Bleves’ lab, are setting-up tools to monitor the assembly of the T6SS MC directly in P. aeruginosa. By breaking the assembly of the T6SS MC, our work will pave the way for the design of more potent drugs that could in turn abolish the ability of P. aeruginosa to colonize immunodepressed patients.

Abdeljalil MADANI : Cyclophostin & Cyclipostins analogs: A new Hope in the fight against M. abscessus

Mycobacterium abscessus is nowadays under the spotlight of the scientific community. This pathogenic mycobacteria is indeed responsible for a wide spectrum of infections involving mostly pulmonary infections in patients with Cystic Fibrosis. M. abscessus is intrinsically resistant to a broad range of antibiotics, including most antitubercular drugs, and is considered the most pathogenic and chemotherapy-resistant rapidly growing mycobacterium. Consequently, with very limited treatment options, the development of new therapeutic approaches to fight this pathogen are urgently needed.

In this context, we have recently synthesized new analogues of Cyclipostins & Cyclophostin (CyCs), compounds naturally produced by Streptomyces species, and evaluated their antibacterial activities against pathogenic mycobacteria (M. marinum, M. bovis BCG and M. tuberculosis), and clinical isolates responsible for nosocomial infections, including Gram-negative and Gram-positive bacteria as well as rapidly growing mycobacteria (RGM) belonging to the M. chelonae-abscessus clade.

Remarkably, the CyCs displayed very low toxicity towards host cells and their inhibitory activity was exclusively restricted to mycobacteria. The best candidate, CyC17, showed a high selectivity for mycobacteria with MIC values (<2 up to 40 µg/mL) comparable to those of most classical antibiotics used to treat M. abscessus infections. Of importance, several CyCs were active against extracellular M. abscessus growth (i.e., CyC17 / CyC18β / CyC25 / CyC26) or against intracellular mycobacteria inside macrophages (i.e., CyC7(α,β) / CyC8(α,β)) with MIC values similar to or better than those of standard antibiotics. An activity-based protein profiling combined with mass spectrometry allowed identification of the potential target enzymes of CyC17/CyC26, mostly being involved in lipid metabolism and/or in cell wall biosynthesis.

Taken together, all these results suggest that these CyC analogs represent a novel class of potent and selective multi-target inhibitor against mycobacteria, and especially against M. abscessus the most drug-resistant mycobacterial species.

David LALAOUNA : The double life of bacterial RNA transcripts

Discovered in the early 80’s, small regulatory RNAs are depicted as key actors in virtually all aspect of bacterial physiology (e.g. primary or secondary metabolism, cell division, virulence). These 50 to 300 nt-long RNA fragments are mostly involved in the post-transcriptional regulation of a subset of messenger RNAs forming their interactomes.

The definition of bacterial sRNAs is still in evolution. For example, they have first been characterized as originating from intergenic regions (chromosome and plasmids). However, recent high-throughput screenings revealed that other loci, such as untranslated regions or transcribed spacers of RNA transcripts, also serve as a source of functional RNA fragments

Matteo BRILLI : Towards Evolutionary Systems Biology

Before DNA sequencing became as affordable as it is today, evolution was mainly studied carrying out detailed comparisons of morphological characteristics of more or less related organisms. Starting in the '70s, sequencing of well-defined genome regions such as the 16S gene enabled molecular phylogenetic analyses, that allowed to better understand (and sometimes confound) the branching patterns in the universal tree of life. Starting in the '90s and up to the present, we enter what we may call the Genome Era, where we are able to compare organisms by looking at the many different information that we obtain from their genome sequence e.g. metabolic abilities, regulatory interactions, sensing systems, virulence factors and antibiotic resistance determinants - even if very often we do not grasp the entire picture of how they work or interact. Additional -omics analysis such as RNA-seq, sRNA-seq, PARE, ChIP-Seq, CLIP-seq etc can help defining regulatory interactions and dynamics.

Camille ANDRIEU : Involvement of Methionine Sulfoxide Reductases in HOCl resistance during host-pathogen interaction

During infection by pathogenic microorganism, oxidative stress is produce by immune cells likemacrophages or neutrophils as a first defence against pathogen. Neutrophils have a specific enzyme,myeloperoxidase, which is able to produce highly oxidative molecules, the hypochlorite acid (HOCl). HOCl is able to disrupt several molecules and especially it oxidizes methionine into methioninesulfoxide. The methionine oxidation is reversible ; this reaction is catalysed by an ubiquitous enzymefamily, the methionine sulfoxide reductase (MSR). We wonder what is the involvement of MSR in HOCl resistance and therefore in bacterial pathogenicity. For this purpose, we used Salmonella typhimurium as a model. A collection of MSR mutants of Salmonella combining different deletion was generated and tested for their sensitivity to industrial HOCl and to an HOCl production system based on purified myeloperoxidases. Our first result showed that a mutant deleted of the five MSR in Salmonella is more sensitive than the wild type strain to HOCl in both conditions. In this way, the phenotypic test and HOCl production by myeloperoxidase provide tools that allowed us to highlight the involvement of MSR in HOCl resistance. Moreover, these results encourage us to consider that MSR could be involved in the infectious power of Salmonella typhimur

Filipe CABREIRO : Host-Microbe-Nutrient Interactions: Metabolic cross-talk for Cancer and Ageing

In the past century, attempts to understand human disease were focused on identifying mutations in the genome responsible or associated with a disease state. Despite great advances in our understanding of disease from this genomic-centric approach, we still do not fully comprehend why similar mutations can lead to a wide-range of disease manifestations. Recent evidence shows that disease states arise from the complex interactions between the genetic make-up of the host and its environment. Nutrition and the microbiome are key environmental factors regulating host physiology but studying these in the context of drug efficacy remains a great challenge. Combining two tractable genetic models, the bacterium E. coli and the nematode C.elegans, we are currently unravelling the complexity underlying such interactions in the efficacy of fluoropyrimidine anticancer drugs 1 and the anti-diabetic drug metformin 2,3 ,4 . Currently, using a 4-way drug-microbe-nutrient-host high-throughput screening approach combined with multi-omics at the host and microbe level (the holobiont) we find that the microbiota integrates nutrition and drug cues through complex signaling networks to drive unique phenotypical outputs in the host. Health benefits to the host conferred by the impact of drugs on the microbiota can be recapitulated through targeted genetic manipulation of signaling or metabolic pathways in bacteria. Further, genetic or pharmacological interventions targeting the microbiota improve host health in other animal models such as Drosophila and mice. Importantly, metabolic modeling of the microbiota in metformin-treated type-2 diabetic patients supports our main findings. Overall, our data shows that the mechanistic understanding of the effects of diet, drugs and intestinal microbiota on host physiology allows their manipulation and may improve health in humans.

Erin GOLEY : Cell division in bacteria: lessons from Caulobacter

Cell division, or cytokinesis, is a fundamental part of bacterial reproduction. Bacterial cell division requires a dedicated protein machine, called the “divisome”, to constrict and remodel the cell envelope and ultimately split one cell physically into two. At the core of the divisome is a polymerizing GTPase called FtsZ that serves to mark the division site, recruit other divisome proteins, and regulate the constriction process by directing cell wall synthesis. The molecular mechanisms by which FtsZ orchestrates cell division remain unclear. Work in my laboratory in recent years using the model Gram-negative alphaproteobacterium Caulobacter crescentus has identified key mechanisms by which FtsZ assembly and function is regulated to promote division. Importantly, our work on multiple regulators of FtsZ assembly and function has converged on the common theme that the ultimate function of FtsZ is to regulate cell wall metabolism. Collectively our findings illuminate molecular details by which FtsZ signals to cell wall metabolic enzymes and support a model for FtsZ as a “dynamic activator” of the cell wall synthesis that drives division.

Illumina : Applications of Next Generation Sequencing in Microbiology

Through NGS, we are learning that we have 100 trillion friends on us which can promote health, – the microbiome.

NGS is enabling us to move beyond benchmarking one genome or a set of known markers to comparing samples to with a multitude of genomes and unlocking the microbiome (our 2nd genome) to promote human health.

First, we are going to show how NGS is enabling researchers to detect pathogens and control outbreaks in food, livestock, and the environment to controlling the transmission of deadly antimicrobial resistance in healthcare settings – core to these applications and getting global precedent is controlling the spread of antimicrobial resistance. Second, we are going to show how Illumina’s solutions are enabling scientists to make groundbreaking, novel discoveries that impacts health and wellness with the community of microbes inside and all around us.

Sam ILLINGWORTH : Facilitating Dialogue Though Creativity

Abstract: In this keynote presentation Dr Sam Illingworth will discuss how the Arts can be used to facilitate meaningful dialogue between scientists and non-scientists. By presenting a series of case studies from his own work, Dr Illingworth will demonstrate how media such as poetry and games can be used to move away from a deficit model of communication and towards a two-way conversation, in which members of the general public not only learn about scientific research, but contribute towards its development and governance.



Bio: Dr Sam Illingworth is a Senior Lecture in Science Communication at Manchester Metropolitan University, where his research centres around using poetry and games to facilitate dialogue between scientists and non-scientists.

GE Healthcare

Liquid chromatography is the major technique used for proteins and biomolecules purification. This approach is always difficult to follow and time consuming. GE Healthcare Life Sciences has designed a new type of system, easier to use, with a high level of automation allowing more experiments with less time spent in front the system.

AKTATM pure is a flexible and intuitive chromatography system for fast purification of proteins, peptides, and nucleic acids from microgram levels to tens of grams of target product. AKTATM pure is a reliable system where hardware and UNICORN™ software are designed to work together with columns and chromatography media to meet any purification challenge. The system has been developed with 14 different valves type offering a huge capacity of different type of automation. The loop valve is one of the most important options for creating new automation approach. The loop valve support 5 loops for 5 automatic samples injections, for analytical or preparative approach, with a high degree of security. The scientist can run several injections automatically night and day, without any risk of cross-contamination.