8W96

SmChiA with diacetyl chitobiose

8W96 の概要

• エントリーDOI: 10.2210/pdb8w96/pdb
• 分子名称: Chitinase A, 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose, GLYCEROL, ... (4 entities in total)
• 機能のキーワード: product, complex, hydrolase
• 由来する生物種: Serratia marcescens
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 62509.05

構造登録者

Tanaka, Y.,Nakamura, A. (登録日: 2023-09-04, 公開日: 2024-09-04, 最終更新日: 2025-03-19)

主引用文献

Tanaka, Y.,Uchihashi, T.,Nakamura, A.
Product inhibition slow down the moving velocity of processive chitinase and sliding-intermediate state blocks re-binding of product.
Arch.Biochem.Biophys., 752:109854-109854, 2024

PubMed Abstract: Processive movement is the key reaction for crystalline polymer degradation by enzyme. Product release is an important phenomenon in resetting the moving cycle, but how it affects chitinase kinetics was unknown. Therefore, we investigated the effect of diacetyl chitobiose (C2) on the biochemical activity and movement of chitinase A from Serratia marcescens (SmChiA). The apparent inhibition constant of C2 on crystalline chitin degradation of SmChiA was 159 μM. The binding position of C2 obtained by X-ray crystallography was at subsite +1, +2 and Trp275 interact with C2 at subsite +1. This binding state is consistent with the competitive inhibition obtained by biochemical analysis. The apparent inhibition constant of C2 on the moving velocity of high-speed (HS) AFM observations was 330 μM, which is close to the biochemical results, indicating that the main factor in crystalline chitin degradation is also the decrease in degradation activity due to inhibition of processive movement. The Trp275 is a key residue for making a sliding intermediate complex. SmChiA W275A showed weaker activity and affinity than WT against crystalline chitin because it is less processive than WT. In addition, biochemical apparent inhibition constant for C2 of SmChiA W275A was 45.6 μM. W275A mutant showed stronger C2 inhibition than WT even though the C2 binding affinity is weaker than WT. This result indicated that Trp275 is important for the interaction at subsite +1, but also important for making sliding intermediate complex and physically block the rebinding of C2 on the catalytic site for crystalline chitin degradation.

PubMed: 38081338
DOI: 10.1016/j.abb.2023.109854

実験手法

X-RAY DIFFRACTION (2.2 Å)