Chimeragenesis of distantly-related proteins by noncontiguous recombination

Authors

  • Matthew A. Smith,

    1. Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, California 91125
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  • Philip A. Romero,

    1. Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, California 91125
    Current affiliation:
    1. Department of Bioengineering and Therapeutic Sciences, University of California San Francisco, San Francisco, CA 94143
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  • Timothy Wu,

    1. Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, California 91125
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  • Eric M. Brustad,

    1. Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, California 91125
    Current affiliation:
    1. Department of Chemistry, Carolina Center for Genome Sciences, University of North Carolina, Chapel Hill, NC 27599
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  • Frances H. Arnold

    Corresponding author
    1. Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, California 91125
    • Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA 91125
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Abstract

We introduce a method for identifying elements of a protein structure that can be shuffled to make chimeric proteins from two or more homologous parents. Formulating recombination as a graph-partitioning problem allows us to identify noncontiguous segments of the sequence that should be inherited together in the progeny proteins. We demonstrate this noncontiguous recombination approach by constructing a chimera of β-glucosidases from two different kingdoms of life. Although the protein's alpha–beta barrel fold has no obvious subdomains for recombination, noncontiguous SCHEMA recombination generated a functional chimera that takes approximately half its structure from each parent. The X-ray crystal structure shows that the structural blocks that make up the chimera maintain the backbone conformations found in their respective parental structures. Although the chimera has lower β-glucosidase activity than the parent enzymes, the activity was easily recovered by directed evolution. This simple method, which does not rely on detailed atomic models, can be used to design chimeras that take structural, and functional, elements from distantly-related proteins.

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