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The effect of N‐terminal acetylation on the structure of an N‐terminal tropomyosin peptide and αα‐tropomyosin
Author(s) -
Greenfield Norma J.,
Stafford Walter E,
HitchcockDegregori Sarah E.
Publication year - 1994
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1002/pro.5560030304
Subject(s) - peptide , chemistry , tropomyosin , acetylation , sedimentation equilibrium , coiled coil , circular dichroism , ionic strength , protein structure , sedimentation coefficient , random coil , denaturation (fissile materials) , folding (dsp implementation) , crystallography , biophysics , biochemistry , ultracentrifuge , aqueous solution , actin , organic chemistry , biology , enzyme , gene , nuclear chemistry , electrical engineering , engineering
We have used a synthetic peptide consisting of the first 30 residues of striated muscle α‐tropomyosin, with GlyCys added to the C‐terminus, to investigate the effect of N‐terminal acetylation on the conformation and stability of the N‐terminal domain of the coiled‐coil protein. In aqueous buffers at low ionic strength, the reduced, unacetylated 32mer had a very low α‐helical content (approximately 20%) that was only slightly increased by disulfide crosslinking or N‐terminal acetylation. Addition of salt (> 1 M) greatly increased the helical content of the peptide. The CD spectrum, the cooperativity of folding of the peptide, and sedimentation equilibrium ultracentrifugation studies showed that it formed a 2‐chained coiled coil at high ionic strength. Disulfide crosslinking and N‐terminal acetylation both greatly stabilized the coiled‐coil α‐helical conformation in high salt. Addition of ethanol or trifluoroethanol to solutions of the peptide also increased its α‐helical content. However, the CD spectra and unfolding behavior of the peptide showed no evidence of coiled‐coil formation. In the presence of the organic solvents, N‐terminal acetylation had very little effect on the conformation or stability of the peptide. Our results indicate that N‐terminal acetylation stabilizes coiled‐coil formation in the peptide. The effect cannot be explained by interactions with the “helix‐dipole” because the stabilization is observed at very high salt concentrations and is independent of pH. In contrast to the results with the peptide, N‐terminal acetylation has only small effects on the overall stability of tropomyosin.