1YJB

SUBTILISIN BPN' 8397+1 (E.C. 3.4.21.14) (MUTANT WITH MET 50 REPLACED BY PHE, ASN 76 REPLACED BY ASP, GLY 169 REPLACED BY ALA, GLN 206 REPLACED BY CYS, ASN 218 REPLACED BY SER AND LYS 256 REPLACED BY TYR) (M50F, N76D, G169A, Q206C, N218S, AND K256Y) IN 35% DIMETHYLFORMAMIDE

1YJB の概要

• エントリーDOI: 10.2210/pdb1yjb/pdb
• 分子名称: SUBTILISIN 8397+1, CALCIUM ION (3 entities in total)
• 機能のキーワード: hydrolase, sporulation, serine protease, zymogen, hydrolase (serine protease)
• 由来する生物種: Bacillus amyloliquefaciens
• 細胞内の位置: Secreted: P00782
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 27661.65

構造登録者

Kidd, R.D.,Farber, G.K. (登録日: 1996-01-16, 公開日: 1996-07-11, 最終更新日: 2024-10-23)

主引用文献

Kidd, R.D.,Sears, P.,Huang, D.H.,Witte, K.,Wong, C.H.,Farber, G.K.
Breaking the low barrier hydrogen bond in a serine protease.
Protein Sci., 8:410-417, 1999

PubMed Abstract: The serine protease subtilisin BPN' is a useful catalyst for peptide synthesis when dissolved in high concentrations of a water-miscible organic co-solvent such as N,N-dimethylformamide (DMF). However, in 50% DMF, the k(cat) for amide hydrolysis is two orders of magnitude lower than in aqueous solution. Surprisingly, the k(cat) for ester hydrolysis is unchanged in 50% DMF. To explain this alteration in activity, the structure of subtilisin 8397+1 was determined in 20, 35, and 50% (v/v) DMF to 1.8 A resolution. In 50% DMF, the imidazole ring of His64, the central residue of the catalytic triad, has rotated approximately 180 degrees around the Cbeta-Cgamma bond. Two new water molecules in the active site stabilize the rotated conformation. This rotation places His64 in an unfavorable geometry to interact with the other members of the catalytic triad, Ser221 and Asp32. NMR experiments confirm that the characteristic resonance due to the low barrier hydrogen bond between the His64 and Asp32 is absent in 50% DMF. These experiments provide a clear structural basis for the change in activity of serine proteases in organic co-solvents.

PubMed: 10048334

実験手法

X-RAY DIFFRACTION (1.8 Å)