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Design of an allosterically regulated retroaldolase.


ABSTRACT: We employed a minimalist approach for design of an allosterically controlled retroaldolase. Introduction of a single lysine residue into the nonenzymatic protein calmodulin led to a 15,000-fold increase in the second order rate constant for retroaldol reaction with methodol as a substrate. The resulting catalyst AlleyCatR is active enough for subsequent directed evolution in crude cell bacterial lysates. AlleyCatR's activity is allosterically regulated by Ca(2+) ions. No catalysis is observed in the absence of the metal ion. The increase in catalytic activity originates from the hydrophobic interaction of the substrate (∼2000-fold) and the change in the apparent pKa of the active lysine residue.

SUBMITTER: Raymond EA 

PROVIDER: S-EPMC4380986 | BioStudies | 2015-01-01

SECONDARY ACCESSION(S): 1CLL

REPOSITORIES: biostudies

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