Marie-Claude SerreFrench National Centre for Scientific Research | CNRS · Institut de Biologie Intégrative de la Cellule
Marie-Claude Serre
PhD Microbiology
About
37
Publications
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Introduction
My different projects are based upon the study of the SSV1 viral cycle. This fusellovirus infects thermoacidophilic archaea of the Sulfolobus genera. By using different approaches (molecular biology, biochemistry, electron microscopy) we want to identify the function of several proteins of unknown function that are conserved in all SSVs to describe at the molecular level the main stages of SSV1 viral cycle.
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Additional affiliations
October 2017 - present
Institut de Biologie Intégrative de la Cellule
Position
- Communication Officer
January 2015 - September 2017
January 2014 - December 2014
Education
July 2004 - October 2005
September 1986 - December 1989
September 1984 - September 1986
Publications
Publications (37)
Gene transfer allows transient or permanent genetic modifications of cells for experimental or therapeutic purposes. Gene delivery by HIV-derived lentiviral vector (LV) is highly effective but the risk of insertional mutagenesis is important and the random/uncontrollable integration of the DNA vector can deregulate the cell transcriptional activity...
Tyrosine recombinases are conserved in the three kingdoms of life. Here we present the first crystal structure of a full-length archaeal tyrosine recombinase, XerA from Pyrococcus abyssi, at 3.0 Å resolution. In the absence of DNA substrate XerA crystallizes as a dimer where each monomer displays a tertiary structure similar to that of DNA-bound Ty...
Organisation of the XerA active site.
A. The structure of the active sites within the XerA dimer is at the top, with catalytic residue side chains in orange. One of the two catalytic Tyr is in red. cis and trans active site organisations are cartooned for wild type and mutants used in complementation assays. Proficient active sites are indicated by...
List of protein structures discussed in the article.
(DOC)
Residues involved in the XerA dimer interface. The 2 monomers of both XerA and Cre are in green and grey respectively. A. Three 90° rotation views of the XerA dimer. The interaction surfaces are respectively in dark green and black for the green and grey monomers. A close-up of the C-terminal interaction surface is presented in two orientations and...
Sedimentation velocity analysis of XerA at 25 °C. Detection of the protein concentration as a function of radial position and time was performed by optical density measurements at a wavelength of 290 nm. Main figure: Continuous sedimentation coefficient distribution analysis (inset) Sedimentation characteristic of the monomer, dimer and tetramer fo...
Tyrosine recombinases are conserved in the three kingdoms of life. Here we present the first crystal structure of a full-length archaeal tyrosine recombinase, XerA from Pyrococcus abyssi, at 3.0 Å resolution. In the absence of DNA substrate XerA crystallizes as a dimer where each monomer displays a tertiary structure similar to that of DNA-bound Ty...
Thermococcales predicted dif sites and conserved flanking regions. Alignment of predicted dif sites and conserved flanking sequences. The flanking sequences are approximately 23 bp long and AT rich. A consensus sequence was deduced and is represented as a sequence logo (see [53] in the main text).
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XerA enzymatic properties. A. XerA substrate specificity. The three substrates were incubated for 1 hr at 65°C with or without 10 pmol of XerA. Recombination products are only observed on the pBend2-dif substrate. B: time course of XerA-mediated recombination at 65°C. C: XerA was pre-incubated at different times at 65°C and then mixed with the dif-...
Whole genome alignments of the four Thermococcales genomes. A. Alignment of P. horikoshii (X-axis) and P. abyssi (Y-axis) genomes shows that they share several regions with conserved gene order. dif sites (circle) are located in a relatively well-conserved region at 135° from oriC (triangle) in P. horikoshii and at 142° from oriC in P. abyssi. The...
Binding specificity of P. abyssi XerA to specific and non-specific DNA substrates. 40 pmoles of XerA were incubated with dif-Pab or attP substrates at 20°C with increasing amounts of non specific competitor poly(dIdC)2. Bottom panel: quantification of free and bound DNA as a function of poly(dIdC)2 amount. Plain lines, dif-Pab substrate; dotted lin...
Specific genomic positions of oriC-cdc6, xerA genes and dif sites in Thermococcales genomes.
(0.06 MB PDF)
Xer similarity scores. A. Top five E. coli XerC and XerD matches in complete sequenced archaeal genomes. B. Similarities between XerA from Thermococcales.
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Identification and localization of the M. stadtmanae dif site. A single statistical-significant sequence matching the Thermococcales dif sites was found in M. stadtmanae by using HMM search. The dif candidate localizes at the ASPS skew inversion.
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Alignment of the C-terminal domain of Xer proteins from the XerD, XerC, XerA and XerS subfamilies. Left panel: dif binding motif alignment. The XerA putative dif binding motif show high residues conservation with both XerC and XerD motifs. Thermococcales XerA harbour the XerC ‘XRX’ motif signature. XerS proteins show very few residues conserved, me...
Genomic localization of dif sites and xer genes. ASPS skew graphics from T. sibiricus, T. onnurineus and T. gammatolerans. TGGT is the most skewed sequence (ASPS) for all species. Symbols are as in Figure 2. Genomic coordinates of oriC, dif and xerA genes can be found in Table S1.
(0.11 MB PDF)
Sulfolobales dif sites. By using the methodology described in the main text of this article on S. solfataricus, S. acidocaldarius and S. tokodaii genomes, one single sequence that fits all of the requirements (two inverted repeats separated by a spacer of 4–8 base pairs, highly conserved between the three genomes and located inside intergenic regio...
Homologous recombination events between circular chromosomes, occurring during or after replication, can generate dimers that need to be converted to monomers prior to their segregation at cell division. In Escherichia coli, chromosome dimers are converted to monomers by two paralogous site-specific tyrosine recombinases of the Xer family (XerC/D)....
Reverse gyrase is a unique type IA topoisomerase that is able to introduce positive supercoils into DNA in an ATP-dependent
process. ATP is bound to the helicase-like domain of the enzyme that contains most of the conserved motifs found in helicases
of the SF1 and SF2 superfamilies. In this paper, we have investigated the role of the conserved heli...
The DNA topoisomerases are essential for DNA replication, transcription, recombination, as well as for chromosome compaction and segregation. They may have appeared early during the formation of the modern DNA world. Several families and subfamilies of the two types of DNA topoisomerases (I and II) have been described in the three cellular domains...
A class of enzymes, called DNA topoisomerases, is responsible for controlling the topological state of cellular DNA. Among these, type IA topoisomerases form a vast family that is present in all living organisms, including higher eukaryotes, in which they play important roles in genome stability. The known 3D structures of three of these enzymes in...
L'étude des virus d'archaea, la manière dont ils sont capables d'infecter leurs hôtes et éventuellement de réaliser le transfert de certains gènes est d'intérêt pour mieux comprendre les mécanismes moléculaires qui ont permis le brassage de l'information génétique dans le phylum des archaea. Notre modèle d'étude est le fusellovirus SSV1 qui infecte...
The only tyrosine recombinase so far studied in archaea, the SSV1 integrase, harbors several changes in the canonical residues forming the catalytic pocket of this family of recombinases. This raised the possibility of a different mechanism for archaeal tyrosine recombinase. The residues of Int(SSV) tentatively involved in catalysis were modified b...
The transposase (InsAB′) of the insertion element IS1 can create breaks in DNA that lead to induction of the SOS response. We have used the SOS response to InsAB′ to screen for
host mutations that affect InsAB′ function and thus point to host functions that contribute to the IS1 transposition mechanism. Mutations in the hns gene, which codes for a...
DNA transactions such as replication, recombination, and transcription produce various changes in the structure and topology of the DNA molecule. These changes are triggered by specialized enzymes that produce transient or permanent breaks in the DNA backbone and are classified as topoisomerases and recombinases. The role of these enzymes is especi...
SSV1 is a virus infecting the extremely thermophilic archaeon Sulfolobus shibatae. The viral-encoded integrase is responsible for site-specific integration of SSV1 into its host genome. The recombinant enzyme
was expressed inEscherichia coli, purified to homogeneity, and its biochemical properties investigated in vitro. We show that the SSV1 integr...
Inspection of the primary sequence of the IS1 transposase suggested that it carries residues which are characteristic of the active site of integrases of the bacteriophage lambda family (Int). In particular, these include a highly conserved triad: His-Arg-Tyr. The properties of mutants made at each of these positions were investigated in vivo. The...
Each strand transfer catalyzed by the Flp recombinase is the composite of two transesterification reactions. The active nucleophilic species in the two reactions are the catalytic site tyrosine (Tyr-343) of Flp and the 5'-hydroxyl from the Flp-nicked DNA substrate, respectively. A "half recombination site" is capable of undergoing this pair of tran...
The site-specific recombinases Flp and R from Saccharomyces cerevisiae and Zygosaccharomyces rouxii, respectively, are related proteins that share approximately 30% amino acid matches. They exhibit a common reaction mechanism that appears to be conserved within the larger Integrase family of site-specific recombinases. Two regions of the proteins,...
The Flp recombinase of Saccharomyces cerevisae and the related R recombinase of Zygosaccharomyces rouxii can efficiently catalyze strand cleavage and strand exchange reactions in half recombination sites. A half-site consists of one recombinase binding element, a recombinase cleavage site on one strand and a 5' spacer hydroxyl group on the other th...
The Flp recombinase of Saccharomyces cerevisae can mediate strand transfer within a half-site, between two half-sites and between a half-site and a full-site. The ability of "step-arrest" mutants of Flp to partake in half-site reactions has been examined. Arg308 variants of Flp, which show little or no strand cleavage in reactions with normal full-...
The leucine residue at position 178 in the allosteric phosphofructokinase from Escherichia coli has been changed into a tryptophan residue by oligonucleotide-directed mutagenesis. The modified enzyme has been purified to homogeneity, and its enzymatic properties show that this single mutation suppresses the heterotropic interactions without affecti...
The leucine residue at position 178 in the major allosteric phosphofructokinase from Escherichia coli has been replaced by a tryptophan using site-directed mutagenesis. Transformation by the mutated gene of pfk- bacteria results into the expression of a pfk+ phenotype and the production of an active enzyme. The modified protein has been purified an...
In order to investigate the role of the carboxy-terminal segment in the catalytic, regulatory, and structural properties of the major allosteric phosphofructokinase (ATP:D-fructose-6-phosphate-1-phosphotransferase: EC 2.7.1.11) from Escherichia coli, the corresponding gene has been modified at either of two sites using oligonucleotide-directed muta...
Enzymatic properties, renaturation and metabolic role of mannitol-1-phosphate dehydrogenase from Escherichia coli. D-mannitol-1-phosphate dehydrogenase was purified to homogeneity from Escherichia coli, and its physicochemical and enzymatic properties were investigated. The molecular weight of the polypeptide chain is 45,000 as determined by polyac...
The complete nucleotide sequences of the Salmonella typhimurium LT2 and Shigella flexneri 2B crp genes were determined and compared with those of the Escherichia coli K-12 crp gene. The Shigella flexneri gene was almost like the E. coli crp gene, with only four silent base pair changes. The S. typhimurium and E. coli crp genes presented a higher de...
Questions
Questions (3)
I want to check if some proteins have associated lipids by SDS-PAGE analysis (either tris-glycine or tris-tricine gel systems). I am looking for a protocol using black sudan. Has someone already used such a stain? Is there another way to proceed?
I am looking for a protocol using pentylamine to prepare grids for DNA spreading. I cannot access the original reference describing this technique (A new preparation method for dark-field electron microscopy of biomacromolecules, Dubochet et al, 1971, J Ultrastruct Res, 35: 147-167). Does someone has experience with it, and if so, could you provide me with the protocol ?
Thanks for your help.
I work on an hyperthermophilic enveloped archaeal virus. We are starting cryo-EM and the quality of our viral preps is not good enough. I am therefore looking for a good protocol to purify enveloped viruses. All suggestions are welcome.
