Qian M, Haser R, Payan F
LCCMB-CNRS, URA 1296, Université Aix-Marseille II, Faculté de Médecine Nord Bd Pierre Dramard, France.
Protein Sci. 1995 Apr;4(4):747-55. doi: 10.1002/pro.5560040414.
The X-ray structure analysis of a crystal of pig pancreatic alpha-amylase (PPA, EC 3.2.1.1.) that was soaked with the substrate maltopentaose showed electron density corresponding to two independent carbohydrate recognition sites on the surface of the molecule. Both binding sites are distinct from the active site described in detail in our previous high-resolution study of a complex between PPA and a carbohydrate inhibitor (Qian M, Buisson G, Duée E, Haser H, Payan F, 1994, Biochemistry 33:6284-6294). One of the binding sites previously identified in a 5-A-resolution electron density map, lies at a distance of 20 A from the active site cleft and can accommodate two glucose units. The second affinity site for sugar units is located close to the calcium binding site. The crystal structure of the maltopentaose complex was refined at 2.1 A resolution, to an R-factor of 17.5%, with an RMS deviation in bond distances of 0.007 A. The model includes all 496 residues of the enzyme, 1 calcium ion, 1 chloride ion, 425 water molecules, and 3 bound sugar rings. The binding sites are characterized and described in detail. The present complex structure provides the evidence of an increased stability of the structure upon interaction with the substrate and allows identification of an N-terminal pyrrolidonecarboxylic acid in PPA.
用麦芽五糖浸泡猪胰α-淀粉酶（PPA，EC 3.2.1.1.）晶体进行的X射线结构分析显示，在分子表面有对应于两个独立碳水化合物识别位点的电子密度。这两个结合位点均不同于我们先前对PPA与碳水化合物抑制剂复合物进行的高分辨率研究（钱M，比松G，迪厄E，哈泽尔H，帕扬F，1994年，《生物化学》33:6284 - 6294）中详细描述的活性位点。先前在5埃分辨率电子密度图中确定的一个结合位点，位于距活性位点裂隙20埃处，可容纳两个葡萄糖单元。糖单元的第二个亲和位点靠近钙结合位点。麦芽五糖复合物的晶体结构在2.1埃分辨率下进行了精修，R因子为17.5%，键长的均方根偏差为0.007埃。该模型包括酶的所有496个残基、1个钙离子、1个氯离子、425个水分子和3个结合的糖环。对结合位点进行了详细表征和描述。目前的复合物结构提供了与底物相互作用时结构稳定性增加的证据，并有助于鉴定PPA中的N端吡咯烷羧酸。
