NDLI: Compensatory stabilizing role of surface mutations during the directed evolution of dienelactone hydrolase for enhanced activity.

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Compensatory stabilizing role of surface mutations during the directed evolution of dienelactone hydrolase for enhanced activity.

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Compensatory stabilizing role of surface mutations during the directed evolution of dienelactone hydrolase for enhanced activity.

Content Provider	World Health Organization (WHO)-Global Index Medicus
Author	Porter, Joanne L. Collyer, Charles A. Ollis, David L.
Description	Author Affiliation: Porter JL ( Research School of Chemistry, Australian National University, Canberra, ACT, 2601, Australia, jo.porter@anu.edu.au.)
Abstract	Directed evolution is a common tool employed to generate enzymes suitable for industrial use. High thermal stability is often advantageous or even a requirement for biocatalysts, as such the evolution of protein stability is of practical as well as academic interest. Even when evolving enzymes for new or improved catalytic functions, stability is an important factor since it can limit the accumulation rate and number of desired active site mutations. Dienelactone hydrolase, a small monomeric protein, has been previously evolved via a three-stage process to possess enhanced activity and specificity toward non-physiological substrates. In addition to seven active site mutations there were three surface mutations that were thought to increase the stability of the enzyme and compensate for the destabilizing active site mutations. Here, the individual influence of the three surface mutations--Q110L, Y137C and N154D--on the thermal and chemical stability of DLH has been assessed. While the Q110L and N154D mutations improved the thermal stability, the influence of the Y137C mutation was more complex. Individually it was destabilizing both thermally and chemically, but when in the presence of the Q110L and N154D mutations its effect was neutralized in relation to thermal but not chemical stability. In the context of a directed evolution experiment, these compensatory surface mutations play important roles. However, our results show that detrimental mutations can arise, thus the simultaneous monitoring of stability changes while evolving enzymes for enhanced catalytic properties can be beneficial.
File Format	HTM / HTML
ISSN	15723887
Issue Number	1
Volume Number	34
e-ISSN	15734943
Journal	The Protein Journal
Language	English
Publisher	Springer
Publisher Date	2015-02-01
Publisher Place	Netherlands
Access Restriction	One Nation One Subscription (ONOS)
Subject Keyword	Discipline Biochemistry Amino Acid Substitution Carboxylic Ester Hydrolases Chemistry Genetics Directed Molecular Evolution Mutation, Missense Enzyme Stability Protein Structure, Secondary Journal Article Research Support, Non-u.s. Gov't
Content Type	Text
Resource Type	Article
Subject	Organic Chemistry Analytical Chemistry Biochemistry Bioengineering

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