1QZ3

CRYSTAL STRUCTURE OF MUTANT M211S/R215L OF CARBOXYLESTERASE EST2 COMPLEXED WITH HEXADECANESULFONATE

Summary for 1QZ3

• Entry DOI: 10.2210/pdb1qz3/pdb
• Related: 1evq
• Descriptor: CARBOXYLESTERASE EST2, 1-HEXADECANOSULFONIC ACID (3 entities in total)
• Functional Keywords: alpha/beta hydrolase fold, hydrolase
• Biological source: Alicyclobacillus acidocaldarius
• Total number of polymer chains: 1
• Total formula weight: 34557.32

Authors

De Simone, G.,Mandrich, L.,Menchise, V.,Giordano, V.,Febbraio, F.,Rossi, M.,Pedone, C.,Manco, G. (deposition date: 2003-09-15, release date: 2004-03-23, Last modification date: 2024-10-30)

Primary citation

De Simone, G.,Mandrich, L.,Menchise, V.,Giordano, V.,Febbraio, F.,Rossi, M.,Pedone, C.,Manco, G.
A substrate-induced switch in the reaction mechanism of a thermophilic esterase: kinetic evidences and structural basis.
J.Biol.Chem., 279:6815-6823, 2004

PubMed Abstract: The reaction mechanism of the esterase 2 (EST2) from Alicyclobacillus acidocaldarius was studied at the kinetic and structural level to shed light on the mechanism of activity and substrate specificity increase previously observed in its double mutant M211S/R215L. In particular, the values of kinetic constants (k1, k(-1), k2, and k3) along with activation energies (E1, E(-1), E2, and E3) were measured for wild type and mutant enzyme. The previously suggested substrate-induced switch in the reaction mechanism from kcat=k3 with a short acyl chain substrate (p-nitrophenyl hexanoate) to kcat=k2 with a long acyl chain substrate (p-nitrophenyl dodecanoate) was validated. The inhibition afforded by an irreversible inhibitor (1-hexadecanesulfonyl chloride), structurally related to p-nitrophenyl dodecanoate, was studied by kinetic analysis. Moreover the three-dimensional structure of the double mutant bound to this inhibitor was determined, providing essential information on the enzyme mechanism. In fact, structural analysis explained the observed substrate-induced switch because of an inversion in the binding mode of the long acyl chain derivatives with respect to the acyl- and alcohol-binding sites.

PubMed: 14617621
DOI: 10.1074/jbc.M307738200

Experimental method

X-RAY DIFFRACTION (2.3 Å)