Mullins Elwood A, Shi Rongxin, Kotsch Lyle A, Eichman Brandt F
Department of Biological Sciences and Center for Structural Biology, Vanderbilt University, Nashville, Tennessee, United States of America.
PLoS One. 2015 May 15;10(5):e0127733. doi: 10.1371/journal.pone.0127733. eCollection 2015.
DNA glycosylases are important repair enzymes that eliminate a diverse array of aberrant nucleobases from the genomes of all organisms. Individual bacterial species often contain multiple paralogs of a particular glycosylase, yet the molecular and functional distinctions between these paralogs are not well understood. The recently discovered HEAT-like repeat (HLR) DNA glycosylases are distributed across all domains of life and are distinct in their specificity for cationic alkylpurines and mechanism of damage recognition. Here, we describe a number of phylogenetically diverse bacterial species with two orthologs of the HLR DNA glycosylase AlkD. One ortholog, which we designate AlkD2, is substantially less conserved. The crystal structure of Streptococcus mutans AlkD2 is remarkably similar to AlkD but lacks the only helix present in AlkD that penetrates the DNA minor groove. We show that AlkD2 possesses only weak DNA binding affinity and lacks alkylpurine excision activity. Mutational analysis of residues along this DNA binding helix in AlkD substantially reduced binding affinity for damaged DNA, for the first time revealing the importance of this structural motif for damage recognition by HLR glycosylases.
DNA糖基化酶是重要的修复酶，可从所有生物体的基因组中清除各种异常核碱基。单个细菌物种通常含有特定糖基化酶的多个旁系同源物，但这些旁系同源物之间的分子和功能差异尚不清楚。最近发现的类热重复（HLR）DNA糖基化酶分布于生命的所有领域，在对阳离子烷基嘌呤的特异性和损伤识别机制方面有所不同。在这里，我们描述了一些系统发育上不同的细菌物种，它们具有HLR DNA糖基化酶AlkD的两个直系同源物。其中一个直系同源物，我们命名为AlkD2，其保守性明显较低。变形链球菌AlkD2的晶体结构与AlkD非常相似，但缺少AlkD中唯一穿透DNA小沟的螺旋。我们发现AlkD2仅具有较弱的DNA结合亲和力，并且缺乏烷基嘌呤切除活性。对AlkD中沿该DNA结合螺旋的残基进行突变分析，大大降低了对受损DNA的结合亲和力，首次揭示了该结构基序对HLR糖基化酶损伤识别的重要性。
