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Fluorescence studies with potato carboxypeptidase inhibitor
C A Ghiron1, M R Eftink, J Longworth
1Department of Biochemistry, University of Missouri, Columbia.
Biochimica Et Biophysica Acta
|December 5, 1990
Summary
Potato carboxypeptidase inhibitor (CPI) has surface-exposed tryptophan residues. Fluorescence studies indicate solvent exposure and interactions with disulfide bonds, influencing its triplet state.
Area of Science:
- Biochemistry
- Structural Biology
- Protein Fluorescence Spectroscopy
Background:
- Potato carboxypeptidase inhibitor (CPI) is a globular protein with a known X-ray structure.
- The protein contains two tryptophan residues (W22 and W28), predicted to be surface-exposed.
Purpose of the Study:
- To investigate the biophysical properties of CPI, specifically the environment and dynamics of its tryptophan residues.
- To understand the fluorescence characteristics and triplet state behavior of CPI.
Main Methods:
- Fluorescence spectroscopy, including acrylamide and KI quenching experiments.
- Fluorescence lifetime and anisotropy decay measurements.
- Low-temperature phosphorescence studies.
Main Results:
- CPI fluorescence spectrum peaks at 344 nm, indicating solvent-exposed tryptophan residues.
- Stern-Volmer plots suggest significant solvent accessibility.
- Fluorescence lifetime data fit a biexponential decay, and anisotropy decay indicates a rotational correlation time of 1.2 ns.
- Low-temperature phosphorescence reveals a biexponential decay, suggesting perturbation of the tryptophan triplet state.
Conclusions:
- The fluorescence properties of CPI are consistent with solvent-exposed tryptophan residues.
- The rotational dynamics may be influenced by factors beyond global rotation.
- Interactions with nearby disulfide bonds likely perturb the triplet state of at least one tryptophan residue.