Functional properties of the thermostable mutL from Thermotoga maritima

Kim, Tae Kyun; Heo, Seong Dal; Ku, Ja Kang; Ban, Chang Ill

doi:10.5483/bmbrep.2009.42.1.053

Cited by 3 publications

(5 citation statements)

References 17 publications

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“…In order to understand the ATP-dependent functional cycles of MutL endonucleases, structural analyses on the full-length MutL homolgoues should be necessary. Since MutL endonucleases from several thermophilic bacteria have relatively short linker region between N-terminal and C-terminal domains [34, 36, 92, 105], these proteins are expected to be suitable for crystallographic analyses. However, we should not underestimate the importance of this interdomain linker region.…”

Section: Discussionmentioning

confidence: 99%

“…As mentioned above, recent biochemical studies on the endonuclease activity of MutL homologues have been achieved by using homologues from thermophilic bacteria, such as T. thermophilus , A. aeolicus , and Thermotoga maritima [34, 36, 87, 92]. Proteins from these bacteria are extremely stable and suitable for physicochemical examinations including crystallographic analysis.…”

Section: Molecular Functions Of Mutl Homologuesmentioning

confidence: 99%

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DNA Mismatch Repair in Eukaryotes and Bacteria

Fukui

2010

Journal of Nucleic Acids

154

130

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DNA mismatch repair (MMR) corrects mismatched base pairs mainly caused by DNA replication errors. The fundamental mechanisms and proteins involved in the early reactions of MMR are highly conserved in almost all organisms ranging from bacteria to human. The significance of this repair system is also indicated by the fact that defects in MMR cause human hereditary nonpolyposis colon cancers as well as sporadic tumors. To date, 2 types of MMRs are known: the human type and Escherichia coli type. The basic features of the former system are expected to be universal among the vast majority of organisms including most bacteria. Here, I review the molecular mechanisms of eukaryotic and bacterial MMR, emphasizing on the similarities between them.

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Section: Discussionmentioning

confidence: 99%

Section: Molecular Functions Of Mutl Homologuesmentioning

confidence: 99%

DNA Mismatch Repair in Eukaryotes and Bacteria

Fukui

2010

Journal of Nucleic Acids

154

130

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“…ATP hydrolysis kinetic data were obtained for various concentrations of TmMutS2 (0.7, 1.2, and 2 µM) with 500 µM ATP. To calculate the specific ATPase activity of TmMutS2, a fluorometric real-time assay was carried out [51]. The reaction was conducted in a solution consisting of 0.1 M KCl, 50 mM Tris-HCl, pH 8.0, 5 mM MgCl 2 , 90 µM NADH, 0.9 mM PEP, 5.3 U/mL pyruvate kinase, 7.5 U/mL lactate dehydrogenase, 0.5 mM ATP, and 400 nM TmL.…”

Section: Methodsmentioning

confidence: 99%

Characterization of Multi-Functional Properties and Conformational Analysis of MutS2 from Thermotoga maritima MSB8

Jeong

Kim

et al. 2012

PLoS ONE

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The MutS2 homologues have received attention because of their unusual activities that differ from those of MutS. In this work, we report on the functional characteristics and conformational diversities of Thermotoga maritima MutS2 (TmMutS2). Various biochemical features of the protein were demonstrated via diverse techniques such as scanning probe microscopy (SPM), ATPase assays, analytical ultracentrifugation, DNA binding assays, size chromatography, and limited proteolytic analysis. Dimeric TmMutS2 showed the temperature-dependent ATPase activity. The non-specific nicking endonuclease activities of TmMutS2 were inactivated in the presence of nonhydrolytic ATP (ADPnP) and enhanced by the addition of TmMutL. In addition, TmMutS2 suppressed the TmRecA-mediated DNA strand exchange reaction in a TmMutL-dependent manner. We also demonstrated that small-angle X-ray scattering (SAXS) analysis of dimeric TmMutS2 exhibited nucleotide- and DNA-dependent conformational transitions. Particularly, TmMutS2-ADPnP showed the most compressed form rather than apo-TmMutS2 and the TmMutS2-ADP complex, in accordance with the results of biochemical assays. In the case of the DNA-binding complexes, the stretched conformation appeared in the TmMutS2-four-way junction (FWJ)-DNA complex. Convergences of biochemical- and SAXS analysis provided abundant information for TmMutS2 and clarified ambiguous experimental results.

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“…TmMutL ATP lid also has an increased predicted content of α-helix. These changes in ATP lid sequences could contribute to the differences with EcNTD observed for PaNTD, and TmMutL [48]. It is interesting to note that the secondary structure prediction algorithm AGADIR predicts the same order in stability of the helix in the ATP lid for the two proteins.…”

Section: Discussionmentioning

confidence: 94%

“…Thermotoga maritima MutL (TmMutL) forms a full dimer in presence of ADP [48]. TmMutL ATP lid also has an increased predicted content of α-helix.…”

Section: Discussionmentioning

confidence: 99%

Analysis of the Interaction Interfaces of the N-Terminal Domain from Pseudomonas aeruginosa MutL

et al. 2013

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Mismatch Repair System corrects mutations arising from DNA replication that escape from DNA polymerase proofreading activity. This system consists of three main proteins, MutS-L-H, responsible for lesion recognition and repair. MutL is a member of GHKL ATPase family and its ATPase cycle has been proposed to modulate MutL activity during the repair process. Pseudomonas aeruginosa MutL (PaMutL) contains an N-terminal (NTD) ATPase domain connected by a linker to a C-terminal (CTD) dimerization domain that possesses metal ion-dependent endonuclease activity. With the aim to identify characteristics that allow the PaMutL NTD allosteric control of CTD endonuclease activity, we used an in silico and experimental approach to determine the interaction surfaces of P. aeruginosa NTD (PaNTD), and compared it with the well characterized Escherichia coli MutL NTD (EcNTD). Molecular dynamics simulations of PaNTD and EcNTD bound to or free of adenosine nucleotides showed that a significant difference exists between the behavior of the EcNTD and PaNTD dimerization interface, particularly in the ATP lid. Structure based simulations of MutL homologues with endonuclease activity were performed that allowed an insight of the dimerization interface behavior in this family of proteins. Our experimental results show that, unlike EcNTD, PaNTD is dimeric in presence of ADP. Simulations in mixed solvent allowed us to identify the PaNTD putative DNA binding patch and a putative interaction patch located opposite to the dimerization face. Structure based simulations of PaNTD dimer in presence of ADP or ATP suggest that nucleotide binding could differentially modulate PaNTD protein-protein interactions. Far western assays performed in presence of ADP or ATP are in agreement with our in silico analysis.

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Functional properties of the thermostable mutL from Thermotoga maritima

Cited by 3 publications

References 17 publications

DNA Mismatch Repair in Eukaryotes and Bacteria

DNA Mismatch Repair in Eukaryotes and Bacteria

Characterization of Multi-Functional Properties and Conformational Analysis of MutS2 from Thermotoga maritima MSB8

Analysis of the Interaction Interfaces of the N-Terminal Domain from Pseudomonas aeruginosa MutL

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