Han S K, Yoon E T, Cho W
Department of Chemistry (M/C 111), University of Illinois at Chicago, 845 West Taylor Street, Chicago, IL 60607-7061, USA.
Biochem J. 1998 Apr 15;331 ( Pt 2)(Pt 2):353-7. doi: 10.1042/bj3310353.
Mammalian secretory class V phospholipase A2 (PLA2) is a newly discovered PLA2 that is implicated in eicosanoid formation in inflammatory cells. As a first step towards understanding the structure, function and regulation of this PLA2, we constructed a bacterial expression vector for human secretory class V PLA2 (hV-PLA2), over-expressed and purified the protein, and determined its physical and kinetic properties. When compared with human class IIa enzyme (hIIa-PLA2), hV-PLA2 has several distinct properties. First, hV-PLA2 can catalyse the hydrolysis of phosphatidylcholine more effectively than hIIa-PLA2 by two orders of magnitude. Secondly, hV-PLA2 has much higher binding affinity and activity for compactly packed phosphatidylcholine bilayers than hIIa-PLA2. Finally, hV-PLA2 has much reduced thermal stability compared with hIIa-PLA2. These data suggest that hV-PLA2 is better suited than hIIa-PLA2 for acting on the outer cellular membrane and liberating arachidonic acid from membrane phospholipids. Also, the unusually low thermal stability of hV-PLA2 might contribute to tighter regulation of its activities in extracellular media.
哺乳动物分泌型V类磷脂酶A2（PLA2）是一种新发现的磷脂酶A2，与炎症细胞中类花生酸的形成有关。作为了解该磷脂酶A2结构、功能和调控的第一步，我们构建了人分泌型V类磷脂酶A2（hV-PLA2）的细菌表达载体，对该蛋白进行了过量表达和纯化，并测定了其物理和动力学性质。与人类IIa类酶（hIIa-PLA2）相比，hV-PLA2具有几个明显的特性。首先，hV-PLA2催化磷脂酰胆碱水解的效率比hIIa-PLA2高两个数量级。其次，hV-PLA2对紧密堆积的磷脂酰胆碱双层的结合亲和力和活性比hIIa-PLA2高得多。最后，与hIIa-PLA2相比，hV-PLA2的热稳定性大大降低。这些数据表明，与hIIa-PLA2相比，hV-PLA2更适合作用于细胞膜外层并从膜磷脂中释放花生四烯酸。此外，hV-PLA2异常低的热稳定性可能有助于在细胞外介质中对其活性进行更严格的调控。
