Protein Engineering, Vol 11, 789-795, Copyright © 1998 by Oxford University Press
T Matsuura, T Yomo, S Trakulnaleamsai, Y Ohashi, K Yamamoto and I Urabe
Catalase I of Bacillus stearothermophilus has high catalatic and low
peroxidatic activities. The mutant from the first random mutant population,
D130N, which has higher peroxidatic and lower catalatic activities than
those exhibited by the wild-type enzyme, was subjected to second random
mutagenesis in observance of the change in reaction specificity. From the
second mutant population, the mutant I108T/D130N/I222T was selected and
examined. The reaction specificity of the purified enzymes revealed that
catalase I being originally 98% catalase and 2% peroxidase was brought to
58% specificity to peroxidase after two-step adaptive walks. From the
statistical analysis of the two random mutant populations, the average
degree of nonadditivity of the mutational effects was estimated to be 0.13
irrespective of the properties of the enzyme. It was demonstrated that the
distribution pattern of a property of the second mutant population can be
predicted well from the data of the first mutant population by taking into
consideration the degree of nonadditivity. The strategy for an efficient
adaptive walk in directed evolution of enzymes through the prediction of
appropriate mutation rate and effective sample size for further mutation
and selection was presented and discussed.
ARTICLES
Nonadditivity of mutational effects on the properties of catalase I and its application to efficient directed evolution
Department of Biotechnology, Graduate School of Engineering, Osaka University, Suita, Japan.
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