ORCID Profile
0000-0003-4898-7871
Current Organisations
University of Sydney
,
CEA
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Publisher: American Society for Microbiology
Date: 28-02-2018
Abstract: Mutator phenotypes have been described in laboratory-evolved bacteria, as well as in natural isolates. Several genes can be impacted, each of them being associated with a typical mutational spectrum. By studying one of the oldest strains available, the ancestral Escherich strain, we were able to identify its mutator status leading to tremendous genetic ersity among the isolates from various collections and allowing us to reconstruct the phylogeographic history of the strain. This mutator phenotype was probably acquired during the storage of the strain, promoting adaptation to a specific environment. Other mutations in rpoS and efflux pump- and porin-encoding genes highlight the acclimatization of the strain through self-preservation and nutritional competence regulation. This strain history can be viewed as unintentional experimental evolution in culture collections all over the word since 1885, mimicking the long-term experimental evolution of E. coli of Lenski et al. (O. Tenaillon, J. E. Barrick, N. Ribeck, D. E. Deatherage, J. L. Blanchard, A. Dasgupta, G. C. Wu, S. Wielgoss, S. Cruveiller, C. Médigue, D. Schneider, and R. E. Lenski, Nature 536:165–170, 2016, 0.1038/nature18959) that shares numerous molecular features.
Publisher: Cold Spring Harbor Laboratory
Date: 15-02-2023
DOI: 10.1101/2023.02.15.528638
Abstract: The growing availability of large genomic datasets presents an opportunity to discover novel metabolic pathways and enzymatic reactions profitable for industrial or synthetic biological applications. Efforts to identify new enzyme functions in this substantial number of sequences cannot be achieved without the help of bioinformatics tools and the development of new strategies. The classical way to assign a function to a gene uses sequence similarity. However, another way is to mine databases to identify conserved gene clusters (i.e. syntenies) as, in prokaryotic genomes, genes involved in the same pathway are frequently encoded in a single locus with an operonic organisation. This Genomic Context (GC) conservation is considered as a reliable indicator of functional relationships, and thus is a promising approach to improve the gene function prediction. Here we present NetSyn (Network Synteny), a tool, which aims to cluster protein sequences according to the similarity of their genomic context rather than their sequence similarity. Starting from a set of protein sequences of interest, NetSyn retrieves neighbouring genes from the corresponding genomes as well as their protein sequence. Homologous protein families are then computed to measure synteny conservation between each pair of input sequences using a GC score. A network is then created where nodes represent the input proteins and edges the fact that two proteins share a common GC. The weight of the edges corresponds to the synteny conservation score. The network is then partitioned into clusters of proteins sharing a high degree of synteny conservation. As a proof of concept, we used NetSyn on two different datasets. The first one is made of homologous sequences of an enzyme family (the BKACE family, previously named DUF849) to ide it into sub-families of specific activities. NetSyn was able to go further by providing additional subfamilies in addition to those previously published. The second dataset corresponds to a set of non-homologous proteins consisting of different Glycosyl Hydrolases (GH) with the aim of interconnecting them and finding conserved operon-like genomic structures. NetSyn was able to detect the locus of Cellvibrio japonicus for the degradation of xyloglucan. It contains three non-homologous GH and was found conserved in fourteen bacterial genomes. NetSyn is able to cluster proteins according to their genomic context which is a way to make functional links between proteins without taking into count their sequence similarity only. We showed that NetSyn is efficient in exploring large protein families to define iso-functional groups. It can also highlight functional interactions between proteins from different families and predicts new conserved genomic structures that have not yet been experimentally characterised. NetSyn can also be useful in pinpointing mis-annotations that have been propagated in databases and in suggesting annotations on proteins currently annotated as “unknown”. NetSyn is freely available at abgem/netsyn .
Publisher: Springer Science and Business Media LLC
Date: 18-03-2019
Publisher: Royal Society of Chemistry (RSC)
Date: 2017
DOI: 10.1039/C6GC02652D
Abstract: Hydroxypyruvate was shown to be a nucleophile for class II pyruvate aldolases isolated from bio ersity, allowing unprecedented stereoselective cross-aldol reactions.
Publisher: Springer Science and Business Media LLC
Date: 08-11-2018
DOI: 10.1038/S41598-018-34795-9
Abstract: Iron(II)/α-ketoacid-dependent oxygenases (αKAOs) are enzymes that catalyze the oxidation of unactivated C-H bonds, mainly through hydroxylation. Among these, those that are active towards amino-acids and their derivatives are grouped in the Clavaminate Synthase Like (CSL) family. CSL enzymes exhibit high regio- and stereoselectivities with strict substrate specificity. This study reports the structural elucidation of two new regio ergent members, KDO1 and KDO5, active towards lysine, and the structural and computational analysis of the whole family through modelling and classification of active sites. The structures of KDO1 and KDO5 in complex with their ligands show that one exact position in the active site controls the regioselectivity of the reaction. Our results suggest that the substrate specificity and high stereoselectivity typical of this family is linked to a lid that closes up in order to form a sub-pocket around the side chain of the substrate. This dynamic lid is found throughout the family with varying sequence and length and is associated with a conserved stable dimeric interface. Results from this study could be a starting-point for exploring the functional ersity of the CSL family and direct in vitro screening in the search for new enzymatic activities.
Publisher: Royal Society of Chemistry (RSC)
Date: 2017
DOI: 10.1039/C6GC02652D
Abstract: Hydroxypyruvate was shown to be a nucleophile for class II pyruvate aldolases isolated from bio ersity, allowing unprecedented stereoselective cross-aldol reactions.
Publisher: MDPI AG
Date: 28-09-2020
DOI: 10.3390/MOLECULES25194459
Abstract: Human group IIA secretory phospholipase A2 (hGIIA) promotes the proliferation of cancer cells, making it a compelling therapeutic target, but it is also significant in other inflammatory conditions. Consequently, suitable inhibitors of hGIIA have always been sought. The activation of phospholipases A2 and the catalysis of glycerophospholipid substrates generally leads to the release of fatty acids such as arachidonic acid (AA) and lysophospholipid, which are then converted to mediator compounds, including prostaglandins, leukotrienes, and the platelet-activating factor. However, this ability of hGIIA to provide AA is not a complete explanation of its biological role in inflammation, as it has now been shown that it also exerts proinflammatory effects by a catalysis-independent mechanism. This mechanism is likely to be highly dependent on key specific molecular interactions, and the full mechanistic descriptions of this remain elusive. The current candidates for the protein partners that may mediate this catalysis-independent mechanism are also introduced in this review. A key discovery has been that selective inhibition of the catalysis-independent activity of hGIIA is achieved with cyclised derivatives of a pentapeptide, FLSYK, derived from the primary sequence of hGIIA. The effects of hGIIA on cell function appear to vary depending on the pathology studied, and so its mechanism of action is complex and context-dependent. This review is comprehensive and covers the most recent developments in the understanding of the many facets of hGIIA function and inhibition and the insight they provide into their clinical application for disease treatment. A cyclic analogue of FLSYK, c2, the most potent analogue known, has now been taken into clinical trials targeting advanced prostate cancer.
Publisher: Springer Science and Business Media LLC
Date: 05-06-2017
Abstract: Experimental validation of enzyme function is crucial for genome interpretation, but it remains challenging because it cannot be scaled up to accommodate the constant accumulation of genome sequences. We tackled this issue for the MetA and MetX enzyme families, phylogenetically unrelated families of acyl-L-homoserine transferases involved in L-methionine biosynthesis. Members of these families are prone to incorrect annotation because MetX and MetA enzymes are assumed to always use acetyl-CoA and succinyl-CoA, respectively. We determined the enzymatic activities of 100 enzymes from erse species, and interpreted the results by structural classification of active sites based on protein structure modeling. We predict that >60% of the 10,000 sequences from these families currently present in databases are incorrectly annotated, and suggest that acetyl-CoA was originally the sole substrate of these isofunctional enzymes, which evolved to use exclusively succinyl-CoA in the most recent bacteria. We also uncovered a ergent subgroup of MetX enzymes in fungi that participate only in L-cysteine biosynthesis as O-succinyl-L-serine transferases.
Publisher: Bioscientifica
Date: 02-2021
DOI: 10.1530/ERC-20-0280
Abstract: Mosaic or somatic EPAS1 mutations are associated with a range of phenotypes including pheochromocytoma and/or paraganglioma (PPGL), polycythemia and somatostatinoma. The pathogenic potential of germline EPAS1 variants however is not well understood. We report a number of germline EPAS1 variants occurring in patients with PPGL, including a novel variant c.739C A (p.Arg247Ser) a previously described variant c.1121T A (p.Phe374Tyr) several rare variants, c.581A G (p.His194Arg), c.2353C A (p.Pro785Thr) and c.2365A G (p.Ile789Val) a common variant c.2296A C (p.Thr766Pro). We performed detailed functional studies to understand their pathogenic role in PPGL. In transient transfection studies, EPAS1 /HIF-2α p.Arg247Ser, p.Phe374Tyr and p.Pro785Thr were all stable in normoxia. In co-immunoprecipitation assays, only the novel variant p.Arg247Ser showed diminished interaction with pVHL. A direct interaction between HIF-2α Arg247 and pVHL was confirmed in structural models. Transactivation was assessed by means of a HRE-containing reporter gene in transiently transfected cells, and significantly higher reporter activity was only observed with EPAS1 /HIF-2α p.Phe374Tyr and p.Pro785Thr. In conclusion, three germline EPAS1 variants (c.739C A (p.Arg247Ser), c.1121T A (p.Phe374Tyr) and c.2353C A (p.Pro785Thr)) all have some functional features in common with somatic activating mutations. Our findings suggest that these three germline variants are hypermorphic alleles that may act as modifiers to the expression of PPGLs.
Publisher: Springer Science and Business Media LLC
Date: 17-11-2013
Abstract: Millions of protein database entries are not assigned reliable functions, preventing the full understanding of chemical ersity in living organisms. Here, we describe an integrated strategy for the discovery of various enzymatic activities catalyzed within protein families of unknown or little known function. This approach relies on the definition of a generic reaction conserved within the family, high-throughput enzymatic screening on representatives, structural and modeling investigations and analysis of genomic and metabolic context. As a proof of principle, we investigated the DUF849 Pfam family and unearthed 14 potential new enzymatic activities, leading to the designation of these proteins as β-keto acid cleavage enzymes. We propose an in vivo role for four enzymatic activities and suggest key residues for guiding further functional annotation. Our results show that the functional ersity within a family may be largely underestimated. The extension of this strategy to other families will improve our knowledge of the enzymatic landscape.
Start Date: 2006
End Date: 2009
Funder: FP6-MOBILITY Marie Curie
View Funded ActivityStart Date: 2003
End Date: 2003
Funder: European Science Foundation
View Funded Activity