1ANB

ANIONIC TRYPSIN MUTANT WITH SER 214 REPLACED BY GLU

1ANB の概要

• エントリーDOI: 10.2210/pdb1anb/pdb
• 分子名称: ANIONIC TRYPSIN, CALCIUM ION, BENZAMIDINE, ... (4 entities in total)
• 機能のキーワード: trypsin, anionic, serine protease, hydrolase
• 由来する生物種: Rattus rattus (black rat)
• 細胞内の位置: Secreted, extracellular space: P00763
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 24137.26

構造登録者

Fletterick, R.J.,Mcgrath, M.E. (登録日: 1994-12-21, 公開日: 1997-04-01, 最終更新日: 2024-10-09)

主引用文献

McGrath, M.E.,Vasquez, J.R.,Craik, C.S.,Yang, A.S.,Honig, B.,Fletterick, R.J.
Perturbing the polar environment of Asp102 in trypsin: consequences of replacing conserved Ser214.
Biochemistry, 31:3059-3064, 1992

PubMed Abstract: Much of the catalytic power of trypsin is derived from the unusual buried, charged side chain of Asp102. A polar cave provides the stabilization for maintaining the buried charge, and it features the conserved amino acid Ser214 adjacent to Asp102. Ser214 has been replaced with Ala, Glu, and Lys in rat anionic trypsin, and the consequences of these changes have been determined. Three-dimensional structures of the Glu and Lys variant trypsins reveal that the new 214 side chains are buried. The 2.2-A crystal structure (R = 0.150) of trypsin S214K shows that Lys214 occupies the position held by Ser214 and a buried water molecule in the buried polar cave. Lys214-N zeta is solvent inaccessible and is less than 5 A from the catalytic Asp102. The side chain of Glu214 (2.8 A, R = 0.168) in trypsin S214E shows two conformations. In the major one, the Glu carboxylate in S214E forms a hydrogen bond with Asp102. Analytical isoelectrofocusing results show that trypsin S214K has a significantly different isoelectric point than trypsin, corresponding to an additional positive charge. The kinetic parameter kcat demonstrates that, compared to trypsin, S214K has 1% of the catalytic activity on a tripeptide amide substrate and S214E is 44% as active. Electrostatic potential calculations provide corroboration of the charge on Lys214 and are consistent with the kinetic results, suggesting that the presence of Lys214 has disturbed the electrostatic potential of Asp102.

PubMed: 1554694
DOI: 10.1021/bi00127a005

実験手法

X-RAY DIFFRACTION (2.8 Å)