Sugio S, Kashima A, Kishimoto K, Peisach D, Petsko G A, Ringe D, Yoshimura T, Esaki N
The Green Cross Corporation, Hirakata, Osaka, Japan.
Protein Eng. 1998 Aug;11(8):613-9. doi: 10.1093/protein/11.8.613.
The leucine-to-alanine mutation at residue 201 of D-amino acid aminotransferase provides a unique enzyme which gradually loses its activity while catalyzing the normal transamination; the co-enzyme form is converted from pyridoxal 5'-phosphate to pyridoxamine 5'-phosphate upon the inactivation [Kishimoto,K., Yoshimura,T., Esaki,N., Sugio,S., Manning,J.M. and Soda,K. (1995) J. Biochem., 117, 691-696]. Crystal structures of both co-enzyme forms of the mutant enzyme have been determined at 2.0 A resolution: they are virtually identical, and are quite similar to that of the wild-type enzyme. Significant differences in both forms of the mutant are localized only on the bound co-enzyme, the side chains of Lys145 and Tyr31, and a water molecule sitting on the putative substrate binding site. Detailed comparisons of the structures of the mutant, together with that of the pyridoxamine-5'-phosphate form of the wild-type enzyme, imply that Leu201 would play a crucial role in the transamination reaction by keeping the pyridoxyl ring in the proper location without disturbing its oscillating motion, although the residue seems to not be especially important for the structural integrity of the enzyme.
D-氨基酸转氨酶201位残基的亮氨酸到丙氨酸突变产生了一种独特的酶，该酶在催化正常转氨作用时会逐渐失去活性；失活时辅酶形式从磷酸吡哆醛转化为磷酸吡哆胺[岸本，K.，吉村，T.，江崎，N.，杉尾，S.，曼宁，J.M.和曾田，K.（1995年）《生物化学杂志》，117，691 - 696]。突变酶两种辅酶形式的晶体结构已在2.0 Å分辨率下确定：它们几乎相同，且与野生型酶的晶体结构非常相似。突变体两种形式的显著差异仅局限于结合的辅酶、赖氨酸145和酪氨酸31的侧链以及位于假定底物结合位点的一个水分子。对突变体结构与野生型酶磷酸吡哆胺 - 5'-磷酸形式结构的详细比较表明，亮氨酸201在转氨反应中可能起着关键作用，它能使吡哆醛环保持在适当位置而不干扰其振荡运动，尽管该残基对酶的结构完整性似乎并非特别重要。
