Steiner R A, Rozeboom H J, de Vries A, Kalk K H, Murshudov G N, Wilson K S, Dijkstra B W
Laboratory of Biophysical Chemistry, Department of Chemistry, University of Groningen, Nijenborgh 4, 9747 AG Groningen, The Netherlands.
Acta Crystallogr D Biol Crystallogr. 2001 Apr;57(Pt 4):516-26. doi: 10.1107/s0907444901002530.
Using synchrotron radiation and a CCD camera, X-ray data have been collected from wild-type bovine pancreatic phospholipase A(2) at 100 K to 0.97 A resolution allowing full anisotropic refinement. The final model has a conventional R factor of 9.44% for all reflections, with a mean standard uncertainty for the positional parameters of 0.031 A as calculated from inversion of the full positional least-squares matrix. At 0.97 A resolution, bovine pancreatic phospholipase A(2) reveals for the first time that its rigid scaffolding does not preclude flexibility, which probably plays an important role in the catalytic process. Functionally important regions (the interfacial binding site and calcium-binding loop) are located at the molecular surface, where conformational variability is more pronounced. A cluster of 2-methyl-2,4-pentanediol molecules is present at the entrance of the hydrophobic channel that leads to the catalytic site and mimics the fatty-acid chains of a substrate analogue. Bovine pancreatic phospholipase A(2) at atomic resolution is compared with previous crystallographic structures and with models derived from nuclear magnetic resonance studies. Given the high structural similarity among extracellular phospholipases A(2) observed so far at lower resolution, the results arising from this structural analysis are expected to be of general validity for this class of enzymes.
利用同步辐射和电荷耦合器件相机，在100 K温度下收集了野生型牛胰磷脂酶A(2)的X射线数据，分辨率达到0.97 Å，从而实现了全各向异性精修。最终模型对所有反射的传统R因子为9.44%，根据全位置最小二乘矩阵的反演计算，位置参数的平均标准不确定度为0.031 Å。在0.97 Å分辨率下，牛胰磷脂酶A(2)首次揭示其刚性支架并不排除灵活性，这可能在催化过程中起重要作用。功能重要区域（界面结合位点和钙结合环）位于分子表面，构象变异性更为明显。在通向催化位点的疏水通道入口处存在一簇2-甲基-2,4-戊二醇分子，它们模拟底物类似物的脂肪酸链。将原子分辨率下的牛胰磷脂酶A(2)与先前的晶体结构以及从核磁共振研究中得到的模型进行了比较。鉴于到目前为止在较低分辨率下观察到的细胞外磷脂酶A(2)之间具有高度的结构相似性，预计这一结构分析结果对于这类酶具有普遍有效性。
