TY  - JOUR
A1  - van Aalten, Daan M. F.
A1  - Komander, David
A1  - Synstad, Bjoenar
A1  - Gaseidnes, Sigrid
A1  - Peter, Martin G.
A1  - Eijsink, Vincent G. H.
T1  - Structural Insights into the catalytic mechanism of a family 18 exochitinase
N2  - Chitinase B (ChiB) from Serratia marcescens is a family 18 exochitinase whose catalytic domain has a TIM-barrel fold with a tunnel-shaped active site. We have solved structures of three ChiB complexes that reveal details of substrate binding, substrateassisted catalysis, and product displacement. The structure of an inactive ChiB mutant (E144Q) complexed with a pentameric substrate (binding in subsites 22 to 13) shows closure of the ''roof'' of the active site tunnel. It also shows that the sugar in the 21 position is distorted to a boat conformation, thus providing structural evidence in support of a previously proposed catalytic mechanism. The structures of the active enzyme complexed to Allosamidin (an analogue of a proposed reaction intermediate) and of the active enzyme soaked with pentameric substrate show events after cleavage of the glycosidic bond. The latter structure shows reopening of the roof of the active site tunnel and enzyme-assisted product displacement in the 11 and 12 sites, allowing a water molecule to approach the reaction center. Catalysis is accompanied by correlated structural changes in the core of the TIM barrel that involve conserved polar residues whose functions were hitherto unknown. These changes simultaneously contribute to stabilization of the reaction intermediate and alternation of the pKa of the catalytic acid during the catalytic cycle.
Y1  - 2002
UR  - https://publishup.uni-potsdam.de/frontdoor/index/index/docId/17547
ER  -