Department of Microbiology, Immunology, and Molecular Genetics, and the Molecular Biology Institute, University of California, and the David Geffen School of Medicine, Los Angeles, CA 90095, USA.
DNA Repair (Amst). 2013 May 1;12(5):334-46. doi: 10.1016/j.dnarep.2013.02.002. Epub 2013 Mar 13.
The most important system for correcting replication errors that survive the built in editing system of DNA polymerase is the mismatch repair (MMR) system. We have identified a novel mutator strain yycJ in Bacillus anthracis. Mutations in the yycJ gene result in a spontaneous mutator phenotype with a mutational frequency and specificity comparable to that of MMR-deficient strains such as those with mutations in mutL or mutS. YycJ was annotated as a metallo-β-lactamase (MβL) super family member with unknown activity. In this study we carried out a biochemical characterization of YycJ and demonstrated that a recombinant YycJ protein possesses a 5'-3' exonuclease activity at the 5' termini and at nicks of double-stranded DNA. This activity requires a divalent metal cofactor Mn2+ and is stimulated by 5'-phosphate ends of duplex DNA. The mutagenesis of conserved amino acid residues revealed that in addition to the five MβL family conserved motifs, YycJ appears to have its specific motifs that can be used to distinguish YycJ from other closely related MβL family members. A phylogenetic survey showed that putative YycJ homologs are present in several bacterial phyla as well as in members of the Methanomicrobiales and Thermoplasmales from Archaea. We propose that YycJ represents a new group of MβL fold exonucleases, which is likely to act in the recognition of MMR entry point and subsequent removal of the mismatched base in certain MutH-less bacterial species.
纠正 DNA 聚合酶内置编辑系统中幸存的复制错误的最重要系统是错配修复（MMR）系统。我们已经在炭疽杆菌中鉴定出一种新型的突变体菌株 yycJ。yycJ 基因突变导致自发突变表型，突变频率和特异性可与 MMR 缺陷菌株（如 mutL 或 mutS 基因突变菌株）相媲美。YycJ 被注释为金属-β-内酰胺酶（MβL）超家族成员，具有未知活性。在这项研究中，我们对 YycJ 进行了生化特性分析，并证明重组 YycJ 蛋白在双链 DNA 的 5'末端和缺口处具有 5'-3'外切核酸酶活性。这种活性需要二价金属辅因子 Mn2+，并且受双链 DNA 5'磷酸末端的刺激。保守氨基酸残基的诱变揭示了除了五个 MβL 家族保守基序外，YycJ 似乎还有其特定的基序，可以将其与其他密切相关的 MβL 家族成员区分开来。系统发育调查显示，推定的 yycJ 同源物存在于几个细菌门中，以及古菌的 Methanomicrobiales 和 Thermoplasmales 成员中。我们提出 YycJ 代表了一种新的 MβL 折叠外切核酸酶组，它可能在某些 MutH 缺失细菌物种中识别 MMR 进入点并随后去除错配碱基中起作用。
