Using magnetic relaxation dispersion (MRD),we have previously shown that the four internal water molecules in bovine pancreatic trypsin inhibitor (BPTI) exchange with bulk water on time scales between 10~(-8) and 10~(-4) s at room temperature.Because this exchange is controlled by the protein structure,internal water molecules can be used to probe rare conformational fluctuations.Here,we report ~2H and ~(17)O MRD data at three temperatures for wild-type BPTI and two BPTI variants where the 14-38 disulfide bond has been cleaved by a double Cys->Ser mutation or by disulfide reduction and carboxamido-methylation.The MRD data show that the internal water molecules are conserved on disulfide cleavage.However,the exchange rate of the water molecule buried near the disulfide bond is enhanced by 2-4 orders of magnitude.The relation of water exchange to other dynamic processes in BPTI is discussed.
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