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Enzyme specificity under dynamic control II: Principal component analysis of -lytic protease using global
 

Summary: Enzyme specificity under dynamic control II: Principal
component analysis of -lytic protease using global
and local solvent boundary conditions
NOBUYUKI OTA AND DAVID A. AGARD
Howard Hughes Medical Institute and Department of Biochemistry and Biophysics, University of California, San
Francisco, San Francisco, California 941430448, USA
(RECEIVED March 1, 2001; FINAL REVISION April 10, 2001; ACCEPTED April 16, 2001)
Abstract
The contributions of conformational dynamics to substrate specificity have been examined by the applica-
tion of principal component analysis to molecular dynamics trajectories of -lytic protease. The wild-type
-lytic protease is highly specific for substrates with small hydrophobic side chains at the specificity pocket,
while the Met190Ala binding pocket mutant has a much broader specificity, actively hydrolyzing sub-
strates ranging from Ala to Phe. Based on a combination of multiconformation analysis of cryo-X-ray
crystallographic data, solution nuclear magnetic resonance (NMR), and normal mode calculations, we had
hypothesized that the large alteration in specificity of the mutant enzyme is mainly attributable to changes
in the dynamic movement of the two walls of the specificity pocket. To test this hypothesis, we performed
a principal component analysis using 1-nanosecond molecular dynamics simulations using either a global
or local solvent boundary condition. The results of this analysis strongly support our hypothesis and verify
the results previously obtained by in vacuo normal mode analysis. We found that the walls of the wild-type
substrate binding pocket move in tandem with one another, causing the pocket size to remain fixed so that

  

Source: Agard, David - Department of Biochemistry and Biophysics, University of California at San Francisco

 

Collections: Biotechnology; Biology and Medicine