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Rescue of conformational dynamics in enzyme catalysis by directed evolution

Rational design and directed evolution have proved to be successful approaches to increase catalytic efficiencies of both natural and artificial enzymes. Protein dynamics is recognized as important, but due to the inherent flexibility of biological macromolecules it is often difficult to distinguish... Full description

Journal Title: Nature Communications 03 April 2018, Vol.9(1)
Main Author: Otten, Renee
Other Authors: Liu, Lin , Kenner, Lillian R , Clarkson, Michael W , Mavor, David , Tawfik, Dan S , Kern, Dorothee , Fraser, James S
Format: Electronic Article Electronic Article
Language: English
Subjects:
ID: ISSN: 2041-1723 ; E-ISSN: 2041-1723 ; DOI: 10.1038/s41467-018-03562-9
Link: https://www.osti.gov/servlets/purl/1499927
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recordid: osti_s1499927
title: Rescue of conformational dynamics in enzyme catalysis by directed evolution
format: Article
creator:
  • Otten, Renee
  • Liu, Lin
  • Kenner, Lillian R
  • Clarkson, Michael W
  • Mavor, David
  • Tawfik, Dan S
  • Kern, Dorothee
  • Fraser, James S
subjects:
  • Basic Biological Sciences
  • Inorganic, Organic, Physical, And Analytical Chemistry
  • Biology
ispartof: Nature Communications, 03 April 2018, Vol.9(1)
description: Rational design and directed evolution have proved to be successful approaches to increase catalytic efficiencies of both natural and artificial enzymes. Protein dynamics is recognized as important, but due to the inherent flexibility of biological macromolecules it is often difficult to distinguish which conformational changes are directly related to function. Here, we use directed evolution on an impaired mutant of the proline isomerase CypA and identify two second-shell mutations that partially restore its catalytic activity. We show both kinetically, using NMR spectroscopy, and structurally, by room-temperature X-ray crystallography, how local perturbations propagate through a large allosteric network to facilitate conformational dynamics. The increased catalysis selected for in the evolutionary screen is correlated with an accelerated interconversion between the two catalytically essential conformational sub-states, which are both captured in the high-resolution X-ray ensembles. Our data provide a glimpse of an evolutionary trajectory and show how subtle changes can fine-tune enzyme function.
language: eng
source:
identifier: ISSN: 2041-1723 ; E-ISSN: 2041-1723 ; DOI: 10.1038/s41467-018-03562-9
fulltext: fulltext
issn:
  • 2041-1723
  • 20411723
url: Link


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titleRescue of conformational dynamics in enzyme catalysis by directed evolution
creatorOtten, Renee ; Liu, Lin ; Kenner, Lillian R ; Clarkson, Michael W ; Mavor, David ; Tawfik, Dan S ; Kern, Dorothee ; Fraser, James S
contributorBrandeis Univ., Waltham, MA (United States) (Corporate Author) ; Univ. of California, San Francisco, Ca (United States) (Corporate Author)
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subjectBasic Biological Sciences ; Inorganic, Organic, Physical, And Analytical Chemistry ; Biology
descriptionRational design and directed evolution have proved to be successful approaches to increase catalytic efficiencies of both natural and artificial enzymes. Protein dynamics is recognized as important, but due to the inherent flexibility of biological macromolecules it is often difficult to distinguish which conformational changes are directly related to function. Here, we use directed evolution on an impaired mutant of the proline isomerase CypA and identify two second-shell mutations that partially restore its catalytic activity. We show both kinetically, using NMR spectroscopy, and structurally, by room-temperature X-ray crystallography, how local perturbations propagate through a large allosteric network to facilitate conformational dynamics. The increased catalysis selected for in the evolutionary screen is correlated with an accelerated interconversion between the two catalytically essential conformational sub-states, which are both captured in the high-resolution X-ray ensembles. Our data provide a glimpse of an evolutionary trajectory and show how subtle changes can fine-tune enzyme function.
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Rational design and directed evolution have proved to be successful approaches to increase catalytic efficiencies of both natural and artificial enzymes. Protein dynamics is recognized as important, but due to the inherent flexibility of biological macromolecules it is often difficult to distinguish which conformational changes are directly related to function. Here, we use directed evolution on an impaired mutant of the proline isomerase CypA and identify two second-shell mutations that partially restore its catalytic activity. We show both kinetically, using NMR spectroscopy, and structurally, by room-temperature X-ray crystallography, how local perturbations propagate through a large allosteric network to facilitate conformational dynamics. The increased catalysis selected for in the evolutionary screen is correlated with an accelerated interconversion between the two catalytically essential conformational sub-states, which are both captured in the high-resolution X-ray ensembles. Our data provide a glimpse of an evolutionary trajectory and show how subtle changes can fine-tune enzyme function.

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Rational design and directed evolution have proved to be successful approaches to increase catalytic efficiencies of both natural and artificial enzymes. Protein dynamics is recognized as important, but due to the inherent flexibility of biological macromolecules it is often difficult to distinguish which conformational changes are directly related to function. Here, we use directed evolution on an impaired mutant of the proline isomerase CypA and identify two second-shell mutations that partially restore its catalytic activity. We show both kinetically, using NMR spectroscopy, and structurally, by room-temperature X-ray crystallography, how local perturbations propagate through a large allosteric network to facilitate conformational dynamics. The increased catalysis selected for in the evolutionary screen is correlated with an accelerated interconversion between the two catalytically essential conformational sub-states, which are both captured in the high-resolution X-ray ensembles. Our data provide a glimpse of an evolutionary trajectory and show how subtle changes can fine-tune enzyme function.

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date2018-04-03