McLeod Neahlanna R, Brolich Matthew A, Damiani Michael J, O'Neill Melanie A
Department of Chemistry, Simon Fraser University, Burnaby, BC V5A1S6, Canada.
Biochem Biophys Res Commun. 2009 Jul 31;385(3):424-9. doi: 10.1016/j.bbrc.2009.05.087. Epub 2009 May 27.
Cryptochromes (CRY) are light-responsive flavoproteins that play central roles in nature and human health, including circadian rhythm regulation. They are closely related to photolyases (PL), but, unlike PL, they cannot repair cyclobutane pyrimidine dimers (CPD) or [6-4] photoproducts in duplex DNA. Yet, if the barrier for flipping the CPD from the duplex is reduced, CRY-DASH, the subclass most structurally homologous to CPD PL, binds and repairs CPD like PL. Here, using limited proteolysis, we have identified the most flexible loops in CPD PL. One corresponds to a "recognition loop" that changes conformation substantially during substrate binding, and engages key interactions with the flipped CPD and the complementary DNA strand. Proteolysis kinetics reveal that the homologous loop in CRY-DASH is at least 10-fold more reactive. We propose that heightened dynamics of the recognition loop in CRY-DASH contribute to its compromised DNA base flipping, and its evolution of divergent function from PL.
隐花色素（CRY）是光响应黄素蛋白，在自然界和人类健康中发挥核心作用，包括昼夜节律调节。它们与光解酶（PL）密切相关，但与PL不同的是，它们不能修复双链DNA中的环丁烷嘧啶二聚体（CPD）或[6-4]光产物。然而，如果将CPD从双链中翻转出来的障碍降低，与CPD PL在结构上最同源的亚类CRY-DASH就会像PL一样结合并修复CPD。在这里，我们通过有限蛋白酶解鉴定了CPD PL中最灵活的环。其中一个对应于一个“识别环”，它在底物结合过程中构象会发生显著变化，并与翻转的CPD和互补DNA链进行关键相互作用。蛋白酶解动力学表明，CRY-DASH中的同源环反应性至少高10倍。我们提出，CRY-DASH中识别环的动态增强导致其受损的DNA碱基翻转，以及其与PL功能分化的进化。
