Structure of the beta 2 homodimer of bacterial luciferase from Vibrio harveyi: X-ray analysis of a kinetic protein folding trap.
Structure of the beta 2 homodimer of bacterial luciferase from Vibrio harveyi: X-ray analysis of a kinetic protein folding trap.
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哈维氏弧菌细菌荧光素酶 β2 同二聚体的结构：动力学蛋白质折叠陷阱的 X 射线分析。
DOI：
10.1002/pro.5560060103
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发表时间：
1997
期刊：
影响因子：
--
通讯作者：
Rayment,I
中科院分区：
文献类型：
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作者：
Thoden,JB;Holden,HM;Fisher,AJ;Sinclair,JF;Wesenberg,G;Baldwin,TO;Rayment,I
从哈维氏弧菌中分离到的荧光素酶是一种Anaβ异源二聚体。当在体外或体内没有α亚基的情况下允许折叠时，酶的β亚基将形成动力学稳定的同源二聚体，即使在PH7.0和18°C的5 M尿素中长时间孵育后也不会展开。这种形式的β亚基，通过折叠途径上的动力学分配而产生，似乎构成了异二聚体酶的动力学捕获替代方案(辛克莱JF，齐格勒MM，鲍德温)。1994年。蛋白质折叠过程中的动力学分配会产生多种天然状态。在这里，我们描述了来自V的荧光素酶的β2同源二聚体的X射线晶体结构。哈维确定并在1.95á分辨率下进行精炼。所用晶体属于正交晶系P212121，晶胞尺寸a=58.8A，b=62.0A，c=218.2，每个不对称单元含有一个二聚体。与在功能性荧光素酶αβ异源二聚体中观察到的一样，每个β亚基的主要三级结构基序由(α/β)8桶(Fisher AJ，Raushel FM，Baldwin to，Rayment I.1995)组成。哈维氏弧菌细菌荧光素酶的三维结构，分辨率为2.4；生化34：6581-6586)。杂二聚体和同二聚体的α亚基之间的β碳坐标的均方根偏差为0.7o。这一高分辨率的X射线分析表明，在β2同源二聚体形成时没有发生“结构域”或“环”交换，因此β2物种对变性的稳定性不能用这样简单的术语来解释。事实上，在β2同二聚体和αβ异二聚体中观察到的亚基：亚基界面在氢键模式和埋藏表面积上非常相似。
Luciferase, as isolated fromVibrio harveyi, is anaβheterodimer. When allowed to fold in the absence of the α subunit, either in vitro or in vivo, the β subunit of the enzyme will form a kinetically stable homodimer that does not unfold even after prolonged incubation in 5 M urea at pH 7.0 and 18 °C. This form of the β subunit, arising via kinetic partitioning on the folding pathway, appears to constitute a kinetically trapped alternative to the heterodimeric enzyme (Sinclair JF, Ziegler MM, Baldwin TO. 1994. Kinetic partitioning during protein folding yields multiple native states.Nature Struct Biol 1: 320–326). Here we describe the X‐ray crystal structure of theβ2homodimer of luciferase fromV. harveyidetermined and refined at 1.95 Å resolution. Crystals employed in the investigation belonged to the orthorhombic space group P212121with unit cell dimensions ofa =58.8 Å,b= 62.0 Å, andc= 218.2 Å and contained one dimer per asymmetric unit. Like that observed in the functional luciferaseαβheterodimer, the major tertiary structural motif of each β subunit consists of an (α/β)8barrel (Fisher AJ, Raushel FM, Baldwin TO, Rayment I. 1995. Three‐dimensional structure of bacterial luciferase fromVibrio harveyiat 2.4 Å resolution.Biochemistry 34: 6581–6586). The root‐mean‐square deviation of the α‐carbon coordinates between the β subunits of the hetero‐ and homodimers is 0.7 Å. This high resolution X‐ray analysis demonstrates that “domain” or “loop” swapping has not occurred upon formation of theβ2homodimer and thus the stability of the β2species to denaturation cannot be explained in such simple terms. In fact, the subunit:subunit interfaces observed in both theβ2homodimer andαβheterodimer are remarkably similar in hydrogen‐bonding patterns and buried surface areas.