NMR Characterization of the Interaction of the Endonuclease Domain of MutL with Divalent Metal Ions and ATP

Mizushima, Ryota; Kim, Ju Yaen; Suetake, Isao; Tanaka, Hiroaki; Takai, Tomoyo; Kamiya, Narutoshi; Takano, Yu; Mishima, Yuichiro; Tsuji, Shoji; Goto, Yuji; Fukui, Kenji; Lee, Young-Ho

doi:10.1371/journal.pone.0098554

Cited by 7 publications

(4 citation statements)

References 42 publications

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“…The endonuclease motifs described above are found in many but not all bacterial MutL proteins ( 3 , 30 ), where they are also believed to comprise Zn 2+ binding endonuclease active sites that are subject to Mn 2+ activation ( 31 ). NMR analysis of the Aquifex aeolicus MutL CTD has shown that Mn 2+ binds in the proximity of the DQHA(X) 2 E(X) 4 E and CPHGRP zinc coordination motifs ( 32 ), which is consistent with our findings based on functional assays.…”

Section: Discussionsupporting

confidence: 90%

The mutagen and carcinogen cadmium is a high-affinity inhibitor of the zinc-dependent MutLα endonuclease

Sherrer

Penland

Modrich

2018

Proc. Natl. Acad. Sci. U.S.A.

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SignificanceMutLα (MLH1-PMS2 heterodimer) is an endonuclease that acts during an early step of eukaryotic mismatch repair. We show that human MutLα endonuclease copurifies with two equivalents of bound zinc, at least one of which resides within the endonuclease active site. We also show that cadmium, a known inhibitor of zinc-dependent enzymes and a potent mutagen and carcinogen, is a high-affinity inhibitor of MutLα endonuclease and that exogenous MutLα significantly reverses the mismatch repair defect in cadmium-treated human cell nuclear extract or nuclear extract prepared from cadmium-treated cells. Because the mutagenic action of cadmium is largely due to the selective inhibition of mismatch repair, these findings suggest that MutLα is a primary cadmium target for mutagenesis and presumably, carcinogenesis as well.

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Section: Discussionsupporting

confidence: 90%

The mutagen and carcinogen cadmium is a high-affinity inhibitor of the zinc-dependent MutLα endonuclease

Sherrer

Penland

Modrich

2018

Proc. Natl. Acad. Sci. U.S.A.

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“…Alternatively, it is possible that the position M3 is the binding site for manganese ion that is critical for the endonuclease activity. Mizushima et al (47) have revealed that, using NMR spectroscopy, some of zinc-coordinating residues also participate in the binding of manganese ion. However, we cannot exclude the possibility that the position M3 does not reflect the physiological metal-binding state because our crystallization buffer used here contained an unphysiological concentration (50 mM) of cadmium ion.…”

Section: Resultsmentioning

confidence: 99%

Structural Features and Functional Dependency on β-Clamp Define Distinct Subfamilies of Bacterial Mismatch Repair Endonuclease MutL

Fukui

Baba

Kumasaka

et al. 2016

Journal of Biological Chemistry

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In early reactions of DNA mismatch repair, MutS recognizes mismatched bases and activates MutL endonuclease to incise the error-containing strand of the duplex. DNA sliding clamp is responsible for directing the MutL-dependent nicking to the newly synthesized/error-containing strand. In Bacillus subtilis MutL, the ␤-clamp-interacting motif (␤ motif) of the C-terminal domain (CTD) is essential for both in vitro direct interaction with ␤-clamp and in vivo repair activity. A large cluster of negatively charged residues on the B. subtilis MutL CTD prevents nonspecific DNA binding until ␤ clamp interaction neutralizes the negative charge. We found that there are some bacterial phyla whose MutL endonucleases lack the ␤ motif. For example, the region corresponding to the ␤ motif is completely missing in Aquifex aeolicus MutL, and critical amino acid residues in the ␤ motif are not conserved in Thermus thermophilus MutL. We then revealed the 1.35 Å-resolution crystal structure of A. aeolicus MutL CTD, which lacks the ␤ motif but retains the metalbinding site for the endonuclease activity. Importantly, there was no negatively charged cluster on its surface. It was confirmed that CTDs of ␤ motif-lacking MutLs, A. aeolicus MutL and T. thermophilus MutL, efficiently incise DNA even in the absence of ␤-clamp and that ␤-clamp shows no detectable enhancing effect on their activity. In contrast, CTD of Streptococcus mutans, a ␤ motif-containing MutL, required ␤-clamp for the digestion of DNA. We propose that MutL endonucleases are divided into three subfamilies on the basis of their structural features and dependence on ␤-clamp.

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“…The C-terminal domains of MutL are constitutively dimerized, whereas the N-terminal domains dimerize only upon ATP binding to form a ring. In organisms with a canonical MMR system, the MutL C-terminal domains possess endonuclease motifs that bind two Zn 2+ ions [49–52] (see minireviews by Kadyrova and Kadyrov and by Groothuizen and Sixma in this issue), and generate single-stranded breaks in DNA [50, 51, 53–56]. In organisms with a methyl-directed MMR system, the C-terminal domains have similar folds, but the endonuclease motifs and metal binding are absent [57]; however, these domains are involved in binding to and activating downstream components of the methyl-directed MMR pathway [58–60].…”

Section: Mmr In E Colimentioning

confidence: 99%

Evolution of the methyl directed mismatch repair system in Escherichia coli

Putnam

2016

DNA Repair

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DNA mismatch repair (MMR) repairs mispaired bases in DNA generated by replication errors. MutS or MutS homologs recognize mispairs and coordinate with MutL or MutL homologs to direct excision of the newly synthesized DNA strand. In most organisms, the signal that discriminates between the newly synthesized and template DNA strands has not been definitively identified. In contrast, Escherichia coli and some related gammaproteobacteria use a highly elaborated methyl-directed MMR system that recognizes Dam methyltransferase modification sites that are transiently unmethylated on the newly synthesized strand after DNA replication. Evolution of methyl-directed MMR is characterized by the acquisition of Dam and the MutH nuclease and by the loss of the MutL endonuclease activity. Methyl-directed MMR is present in a subset of Gammaproteobacteria belonging to the orders Enterobacteriales, Pasteurellales, Vibrionales, Aeromonadales, and a subset of the Alteromonadales (the EPVAA group) as well as in gammaproteobacteria that have obtained these genes by horizontal gene transfer, including the medically relevant bacteria Fluoribacter, Legionella, and Tatlockia and the marine bacteria Methylophaga and Nitrosococcus.

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NMR Characterization of the Interaction of the Endonuclease Domain of MutL with Divalent Metal Ions and ATP

Cited by 7 publications

References 42 publications

The mutagen and carcinogen cadmium is a high-affinity inhibitor of the zinc-dependent MutLα endonuclease

The mutagen and carcinogen cadmium is a high-affinity inhibitor of the zinc-dependent MutLα endonuclease

Structural Features and Functional Dependency on β-Clamp Define Distinct Subfamilies of Bacterial Mismatch Repair Endonuclease MutL

Evolution of the methyl directed mismatch repair system in Escherichia coli

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