June Cynthia M, Muckenthaler Taylor J, Schroder Emma C, Klamer Zachary L, Wawrzak Zdzislaw, Powers Rachel A, Szarecka Agnieszka, Leonard David A
Department of Chemistry, Grand Valley State University, Allendale, Michigan, 49401.
Department of Cell and Molecular Biology, Grand Valley State University, Allendale, Michigan, 49401.
Protein Sci. 2016 Dec;25(12):2152-2163. doi: 10.1002/pro.3040. Epub 2016 Sep 26.
OXA-51 is a class D β-lactamase that is thought to be the native carbapenemase of Acinetobacter baumannii. Many variants of OXA-51 containing active site substitutions have been identified from A. baumannii isolates, and some of these substitutions increase hydrolytic activity toward carbapenem antibiotics. We have determined the high-resolution structures of apo OXA-51 and OXA-51 with one such substitution (I129L) with the carbapenem doripenem trapped in the active site as an acyl-intermediate. The structure shows that acyl-doripenem adopts an orientation very similar to carbapenem ligands observed in the active site of OXA-24/40 (doripenem) and OXA-23 (meropenem). In the OXA-51 variant/doripenem complex, the indole ring of W222 is oriented away from the doripenem binding site, thereby eliminating a clash that is predicted to occur in wildtype OXA-51. Similarly, in the OXA-51 variant complex, L129 adopts a different rotamer compared to I129 in wildtype OXA-51. This alternative position moves its side chain away from the hydroxyethyl moiety of doripenem and relieves another potential clash between the enzyme and carbapenem substrates. Molecular dynamics simulations of OXA-51 and OXA-51 I129L demonstrate that compared to isoleucine, a leucine at this position greatly favors a rotamer that accommodates the ligand. These results provide a molecular justification for how this substitution generates enhanced binding affinity for carbapenems, and therefore helps explain the prevalence of this substitution in clinical OXA-51 variants.
OXA-51是一种D类β-内酰胺酶，被认为是鲍曼不动杆菌的天然碳青霉烯酶。已从鲍曼不动杆菌分离株中鉴定出许多含有活性位点取代的OXA-51变体，其中一些取代增加了对碳青霉烯类抗生素的水解活性。我们已经确定了无配体OXA-51和具有一种此类取代（I129L）的OXA-51的高分辨率结构，其中碳青霉烯类药物多利培南作为酰基中间体被困在活性位点中。该结构表明，酰基多利培南采取的取向与在OXA-24/40（多利培南）和OXA-23（美罗培南）活性位点中观察到的碳青霉烯配体非常相似。在OXA-51变体/多利培南复合物中，W222的吲哚环远离多利培南结合位点，从而消除了预计在野生型OXA-51中发生的冲突。同样，在OXA-51变体复合物中，与野生型OXA-51中的I129相比，L129采取了不同的旋转异构体。这个替代位置使其侧链远离多利培南的羟乙基部分，并缓解了酶与碳青霉烯底物之间的另一个潜在冲突。OXA-51和OXA-51 I129L的分子动力学模拟表明，与异亮氨酸相比，该位置的亮氨酸极大地有利于容纳配体的旋转异构体。这些结果为这种取代如何产生对碳青霉烯类药物增强的结合亲和力提供了分子依据，因此有助于解释这种取代在临床OXA-51变体中的普遍性。
