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| Thebault, Marie-t; Izem, Lahoucine; Leroy, Jean Paul; Gobin, Eric; Charrier, Gregory; Raffin, Jean-paul. |
| AMP-deaminase activity was measured in white muscle from a wide range of fish, including one cyclostome, 13 chondrosteans, and one teleost to elucidate the pattern of the AMP-deaminase activity in white muscle of fish. Compared to a mammalian (rat) muscle extract, low enzyme activities are found in the cyclostome and two elasmobranchs from two families (Scyliorhinidae, Hexanchidae). In contrast, higher AMP-deaminase activities, similar to mammals, are expressed in Squalidae, all families of skates, Chimaeridae and in the teleostean fish. We then compared AMP-deaminase activities in red and white muscles from two representative elasmobranch fish, the dogfish (Scyliorhinus canicula) and the thornback ray (Raja clavata). The fibre type composition and... |
| Tipo: Text |
Palavras-chave: White fibres; Red fibres; Muscle; Histology; Fish; Evolution; Elasmobranch; AMP deaminase. |
| Ano: 2005 |
URL: http://archimer.ifremer.fr/doc/2005/publication-635.pdf |
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| Castrec, Benoit; Rouillon, Christophe; Henneke, Ghislaine; Flament, Didier; Querellou, Joel; Raffin, Jean-paul. |
| Replicative DNA polymerases possess a canonical C-terminal proliferating cell nuclear antigen (PCNA)-binding motif termed the PCNA-interacting protein (PIP) box. We investigated the role of the PIP box on the functional interactions of the two DNA polymerases, PabPol B (family B) and PabPol D (family D), from the hyperthermophilic euryarchaeon Pyrococcus abyssi, with its cognate PCNA. The PIP box was essential for interactions of PabPol B with PCNA, as shown by surface plasmon resonance and primer extension studies. In contrast, binding of PabPol D to PCNA was affected only partially by removing the PIP motif. We identified a second palindromic PIP box motif at the N-terminus of the large subunit of PabPol D that was required for the interactions of PabPol... |
| Tipo: Text |
Palavras-chave: Archaea; PIP box; PCNA binding motifs; DNA polymerases; DNA replication. |
| Ano: 2009 |
URL: http://archimer.ifremer.fr/doc/2009/publication-7317.pdf |
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| Rouillon, Christophe; Henneke, Ghislaine; Flament, Didier; Querellou, Joel; Raffin, Jean-paul. |
| DNA replication in Archaea, as in other organisms, involves large protein complexes called replisomes. In the Euryarchaeota subdomain, only two putative replicases have been identified, and their roles in leading and lagging strand DNA synthesis are still poorly understood. In this study, we focused on the coupling of proliferating cell nuclear antigen (PCNA)loading mechanisms with DNA polymerase function in the Euryarchaea Pyrococcus abyssi. PCNA spontaneously loaded onto primed DNA, and replication factor C dramatically increased this loading. Surprisingly, the family B DNA polymerase (Pol B) also increased PCNA loading, probably by stabilizing the clamp on primed DNA via an essential motif. In contrast, on an RNA-primed DNA template, the PCNA/Pol B... |
| Tipo: Text |
Palavras-chave: RF C; PCNA loading; DNA polymerase switching; DNA replication; Archaea. |
| Ano: 2007 |
URL: http://archimer.ifremer.fr/doc/2007/publication-2638.pdf |
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| Palud, Adeline; Villani, Giuseppe; L'Haridon, Stephane; Querellou, Joel; Raffin, Jean-paul; Henneke, Ghislaine. |
| Spontaneous and induced abasic sites in hyperthermophiles DNA have long been suspected to occur at high frequency. Here, Pyrococcus abyssi was used as an attractive model to analyse the impact of such lesions onto the maintenance of genome integrity. We demonstrated that endogenous AP sites persist at a slightly higher level in P. abyssi genome compared with Escherichia coli. Then, the two replicative DNA polymerases, PabpolB and PabpolD, were characterized in presence of DNA containing abasic sites. Both Pabpols had abortive DNA synthesis upon encountering AP sites. Under running start conditions, PabpolB could incorporate in front of the damage and even replicate to the full-length oligonucleotides containing a specific AP site, but only when present at... |
| Tipo: Text |
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| Ano: 2008 |
URL: http://archimer.ifremer.fr/doc/2008/publication-6113.pdf |
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| Henneke, Ghislaine; Gueguen, Yannick; Flament, Didier; Azam, Philippe; Querellou, Joel; Dietrich, Jacques; Hubscher, Ulrich; Raffin, Jean-paul. |
| The molecular organization of the replication complex in archaea is similar to that in eukaryotes. Only two proteins homologous to subunits of eukaryotic replication factor C (RFC) have been detected in Pyrococcus abyssi (Pab). The genes encoding these two proteins are arranged in tandem. We cloned these two genes and co-expressed the corresponding recombinant proteins in Escherichia coli. Two inteins present in the gene encoding the small subunit (Pab RFC-small) were removed during cloning. The recombinant protein complex was purified by anion-exchange and hydroxyapatite chromatography. Also, the Pab RFC-small subunit could be purified, while the large subunit (Pab RFC-large) alone was completely insoluble. The highly purified Pab RFC complex possessed an... |
| Tipo: Text |
Palavras-chave: PCNA binding domain; Pyrococcus abyssi; Hyperthermophile; Archaea; Replication factor C. |
| Ano: 2002 |
URL: https://archimer.ifremer.fr/doc/2002/publication-675.pdf |
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| Le Breton, Magali; Henneke, Ghislaine; Norais, C; Flament, Didier; Myllykallio, H; Querellou, Joel; Raffin, Jean-paul. |
| We report on the characterization of the DNA primase complex of the hyperthermophilic archaeon Pyrococcus abyssi (Pab). The Pab DNA primase complex is composed of the proteins Pabp41 and Pabp46, which show sequence similarities to the p49 and p58 subunits, respectively, of the eukaryotic polymerase alpha-primase complex. Both subunits were expressed, purified, and characterized. The Pabp41 subunit alone had no RNA synthesis activity but could synthesize long (up to 3 kb) DNA strands. Addition of the Pabp46 subunit increased the rate of DNA synthesis but decreased the length of the DNA fragments synthesized and conferred RNA synthesis capability. Moreover, in our experimental conditions, Pab DNA primase had comparable affinities for ribonucleotides and... |
| Tipo: Text |
Palavras-chave: Strand displacement; Gap filling; DNA primase; Archaea; DNA replication. |
| Ano: 2007 |
URL: http://archimer.ifremer.fr/doc/2007/publication-3520.pdf |
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| Henneke, Ghislaine; Flament, Didier; Hubscher, Ulrich; Querellou, Joel; Raffin, Jean-paul. |
| DNA polymerases carry out DNA synthesis during DNA replication, DNA recombination and DNA repair. During the past five years, the number of DNA polymerases in both eukarya and bacteria has increased to at least 19 and multiple biological roles have been assigned to many DNA polymerases. Archaea, the third domain of life, on the other hand, have only a subset of the eukaryotic-like DNA polymerases. The diversity among the archaeal DNA polymerases poses the intriguing question of their functional tasks. Here, we focus on the two identified DNA polymerases, the family B DNA polymerase B (PabpolB) and the family D DNA polymerase D (PabpolD) from the hyperthermophilic euryarchaeota Pyrococcus abyssi. Our data can be summarized as follows: (i) both Pabpols are... |
| Tipo: Text |
Palavras-chave: DNA polymerase; Strand displacement; Gap filling; Euryarchaea; DNA replication. |
| Ano: 2005 |
URL: http://archimer.ifremer.fr/doc/2005/publication-423.pdf |
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| Henneke, Ghislaine; Raffin, Jean-paul; Ferrari, Elena; Jonsson, Zophonías; Dietrich, Jacques; Hubscher, Ulrich. |
| We have cloned the gene encoding proliferating cell nuclear antigen (PCNA) from the hyperthermophilic euryarchaeote Thermococcus fumicolans (Tfu). Tfu PCNA contains 250 amino acids with a calculated M-r of 28,000 and is 26% identical to human PCNA. Next, Tfu PCNA was overexpressed in Escherichia coli and it showed an apparent molecular mass of 33.5 kDa. The purified Tfu PCNA was tested first with recombinant Tfu DNA polymerase I (Tfu pol) and second with calf thymus DNA polymerase delta (pol delta). When tested with the homologous Tfu pol on bacteriophage lambda DNA, large amounts of Tfu PCNA were required to obtain two- to threefold stimulation. Surprisingly, however, Tfu PCNA was much more efficient than human PCNA in stimulating calf thymus pol delta.... |
| Tipo: Text |
Palavras-chave: DNA polymerase delta; Conservation; Thermococcus fumicolans; Archaea; Hyperthermophile; PCNA. |
| Ano: 2000 |
URL: http://archimer.ifremer.fr/doc/2000/publication-659.pdf |
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