Fast and faster: A designed variant of the B-domain of protein A folds in 3 μsec

Authors

  • Pooja Arora,

    1. Department of Chemistry, Duke University Medical Center, Durham, North Carolina 27710, USA
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  • Terrence G. Oas,

    1. Department of Chemistry, Duke University Medical Center, Durham, North Carolina 27710, USA
    2. Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA
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  • Jeffrey K. Myers

    Corresponding author
    1. Department of Biochemistry, Vanderbilt University Medical Center, Nashville, Tennessee 37232-8725, USA
    • Department of Biochemistry, Vanderbilt University Medical Center, 5140 MRB III, 465 21st Avenue South, Nashville, TN 37232-8725, USA; fax: (615) 936-2211.
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Abstract

We have introduced the mutation glycine 29 to alanine, designed to increase the rate of protein folding, into the B-domain of protein A (BdpA). From NMR lineshape analysis, we find the G29A mutation increases the folding rate constant by threefold; the folding time is 3 μsec. Although wild-type BdpA folds extremely fast, simple-point mutations can still speed up the folding; thus, the folding rate is not evolutionarily maximized. The short folding time of G29A BdpA (the shortest time yet reported) makes it an attractive candidate for an all-atom molecular dynamics simulation that could potentially show a complete folding reaction starting from an extended chain. We also constructed a fluorescent variant of BdpA by mutating phenylalanine 13 to tryptophan, allowing fluorescence-based time-resolved temperature-jump measurements. Temperature jumps and NMR complement each other, and give a very complete picture of the folding kinetics.

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