Bukrinsky J T, Bjerrum M J, Kadziola A
Bioinorganic Group, Chemistry Department, The Royal Veterinary and Agricultural University, Copenhagen, Denmark.
Biochemistry. 1998 Nov 24;37(47):16555-64. doi: 10.1021/bi981678i.
Native carboxypeptidase A has been crystallized in a new crystal form, and the structure has been refined with X-ray data to 2.0 A resolution. In contrast to the previously published structure [Rees, D. C., Lewis, M., and Lipscomb, W. N. (1983) J. Mol. Biol. 168, 367-387], no active-site amino acids are involved in the crystal packing. The important Tyr248 is stabilized inside the active site by a hydrogen bond and by interactions with Ile247. The proposed role of Tyr248 in the induced fit mechanism is therefore not supported by the findings in this structure of native carboxypeptidase A. The structure has a partly populated inhibitory Zn2+ site in close proximity to the catalytic Zn2+ as evident from X-ray anomalous dispersion data. A hydroxo bridge is found between the catalytic Zn2+ and the inhibitory Zn2+ with a Zn2+-Zn2+ distance of 3.48 A. In addition, the inhibitory Zn2+ has Glu270 as a monodentate ligand. No other protein ligands to the inhibitory Zn2+ are seen. The crystals were grown at 0.3 M LiCl and weak evidence for a binding site for partly competitive inhibitory anions is observed.
天然羧肽酶A已结晶成一种新的晶体形式，其结构已利用X射线数据精修至2.0埃分辨率。与之前发表的结构[里斯，D.C.，刘易斯，M.，和利普斯科姆，W.N.（1983年）《分子生物学杂志》168卷，367 - 387页]不同，晶体堆积中不涉及活性位点氨基酸。重要的酪氨酸248通过氢键以及与异亮氨酸247的相互作用稳定在活性位点内部。因此，在天然羧肽酶A的这种结构中所发现的结果不支持酪氨酸248在诱导契合机制中的假定作用。从X射线反常散射数据可以明显看出，该结构在紧邻催化性锌离子处有一个部分占据的抑制性锌离子位点。在催化性锌离子和抑制性锌离子之间发现了一个羟基桥，锌 - 锌距离为3.48埃。此外，抑制性锌离子有谷氨酸270作为单齿配体。未观察到抑制性锌离子的其他蛋白质配体。晶体在0.3 M氯化锂中生长，并且观察到了部分竞争性抑制性阴离子结合位点的微弱证据。
