Evaluation and ranking of enzyme designs

被引:91
作者
Kiss, Gert [1 ]
Roethlisberger, Daniela [2 ]
Baker, David [2 ]
Houk, K. N. [1 ]
机构
[1] Univ Calif Los Angeles, Dept Chem & Biochem, Los Angeles, CA 90095 USA
[2] Univ Washington, Howard Hughes Med Inst, Dept Biochem, Seattle, WA 98195 USA
关键词
Kemp elimination; acid-base catalysis; enzyme active site; proton transfer; molecular dynamics; enzyme design; design refinement; QM/MM; cluster model; PHYSICAL ORGANIC-CHEMISTRY; BASE-CATALYZED DECOMPOSITION; DIELS-ALDER REACTION; PROTON-TRANSFER; ANTIBODY CATALYSIS; KEMP ELIMINATION; COMPUTATIONAL DESIGN; QM/MM SIMULATIONS; HYDROGEN-BONDS; ACTIVE-SITE;
D O I
10.1002/pro.462
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
In 2008, a successful computational design procedure was' reported that yielded active enzyme catalysts for the Kemp elimination. Here, we studied these proteins together with a set of previously unpublished inactive designs to determine the sources of activity or lack thereof, and to predict which of the designed structures are most likely to be catalytic. Methods that range from quantum mechanics (QM) on truncated model systems to the treatment of the full protein with ONIOM QM/MM and AMBER molecular dynamics (MD) were explored. The most effective procedure involved molecular dynamics, and a general MD protocol was established. Substantial deviations from the ideal catalytic geometries were observed for a number of designs. Penetration of water into the catalytic site and insufficient residue-packing around the active site are the main factors that can cause enzyme designs to be inactive. Where in the past, computational evaluations of designed enzymes were too time-extensive for practical considerations, it has now become feasible to rank and refine candidates computationally prior to and in conjunction with experimentation, thus markedly increasing the efficiency of the enzyme design process.
引用
收藏
页码:1760 / 1773
页数:14
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