Fast and faster: a designed variant of the B-domain of protein A folds in 3 microsec |
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Authors: | Arora Pooja Oas Terrence G Myers Jeffrey K |
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Affiliation: | Department of Biochemistry, Vanderbilt University Medical Center, 5140 MRB III, 465 21st Avenue South, Nashville, TN 37232-8725, USA. |
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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 microsec. 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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Keywords: | rapid protein folding mutation denaturant helix propensity dynamic NMR |
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