Kim K P, Han S K, Hong M, Cho W
Department of Chemistry (M/C 111), University of Illinois at Chicago, 845 West Taylor Street, Chicago, IL 60607-7061, USA.
Biochem J. 2000 Jun 15;348 Pt 3(Pt 3):643-7.
Human group-V phospholipase A(2) (hVPLA(2)) is a secretory phospholipase A(2) (PLA(2)) that is involved in eicosanoid formation in such inflammatory cells as macrophages and mast cells. We showed that hVPLA(2) can bind phosphatidylcholine membranes and hydrolyse phosphatidylcholine molecules much more efficiently than human group-IIa PLA(2), which accounts for its high activity on the outer plasma membrane of mammalian cells. To understand the molecular basis of the high phosphatidylcholine specificity of hVPLA(2), we mutated several residues (Gly-53, Glu-56 and Glu-57) that might be involved in interaction with an active-site-bound phospholipid molecule. Phospholipid head-group specificities of mutants determined using polymerized mixed-liposome substrates indicate that a small glycine residue in position 53 is important for accommodating a bulky choline head group. Also, results indicated that two anionic residues, Glu-56 and Glu-57, favourably interact with cationic head groups of phosphatidylcholine and phosphatidylethanolamine. Together, these steric and electrostatic properties of the active site of hVPLA(2) allow for effective binding and hydrolysis of a bulky cationic choline head group of phosphatidylcholine, which is unique among mammalian secretory PLA(2)s.
人类Ⅴ型磷脂酶A2（hVPLA2）是一种分泌型磷脂酶A2（PLA2），参与巨噬细胞和肥大细胞等炎症细胞中类花生酸的形成。我们发现，hVPLA2能够结合磷脂酰胆碱膜并水解磷脂酰胆碱分子，其效率远高于人类Ⅱa型PLA2，这解释了其在哺乳动物细胞外质膜上的高活性。为了解hVPLA2对磷脂酰胆碱高特异性的分子基础，我们对几个可能参与与活性位点结合的磷脂分子相互作用的残基（甘氨酸-53、谷氨酸-56和谷氨酸-57）进行了突变。使用聚合混合脂质体底物测定的突变体的磷脂头部基团特异性表明，53位的一个小甘氨酸残基对于容纳庞大的胆碱头部基团很重要。此外，结果表明，两个阴离子残基谷氨酸-56和谷氨酸-57与磷脂酰胆碱和磷脂酰乙醇胺的阳离子头部基团发生有利的相互作用。总之，hVPLA2活性位点的这些空间和静电特性使得其能够有效结合并水解磷脂酰胆碱的庞大阳离子胆碱头部基团，这在哺乳动物分泌型PLA2中是独一无二的。
