Pig heart calpastatin: identification of repetitive domain structures and anomalous behavior in polyacrylamide gel electrophoresis
- 22 March 1988
- journal article
- research article
- Published by American Chemical Society (ACS) in Biochemistry
- Vol. 27 (6), 1964-1972
- https://doi.org/10.1021/bi00406a024
Abstract
Isolation and nucleotide sequencing of the complementary DNA for pig heart calpastatin have been completed. The amino acid sequence of 713 residues predicted from the nucleotide sequence contains five domains, each composed of approximately 140 amino acid residues. A unique N-terminal domain is followed by four mutually homologous domains. The best fit alignment of these four domains gives residue identities between any two domains of 22.5-36.0%. The analysis of the sequence similarities by several methods also suggests the existence of additional shorter repeats at intervals of 60-80 residues. The calculated molecular weight of pig calpastatin of 713 amino acid residues (Mr 77,122) is significantly lower than the value of purified pig heart calpastatin (Mr 107,000) estimated by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS-PAGE). The expression of the calpastatin genes in Escherichia coli and the detection of the translation products of 713, 366, and 140 amino acid residues by the specific anti-calpastatin antibody indicate that the products always migrate considerably slow on SDS-PAGE, giving an average of 1.53 for the ratio of the molecular weight estimated by SDS-PAGE to the value calculated from the amino acid sequences. It is most likely that the discrepancy in the molecular weight is caused by an anomalous behavior of calpastatin in SDS-PAGE.Keywords
This publication has 20 references indexed in Scilit:
- Molecular diversity of calpastatin in mammalian organsBiochemical and Biophysical Research Communications, 1984
- LARGE-SCALE PURIFICATION OF PORCINE CALPAIN-I AND CALPAIN-II AND COMPARISON OF PROTEOLYTIC FRAGMENTS OF THEIR SUBUNITS1984
- A simple and very efficient method for generating cDNA librariesGene, 1983
- The protein inhibitor of calcium-dependent proteases: Purification from bovine heart and possible mechanisms of regulationArchives of Biochemistry and Biophysics, 1983
- Correspondence of homologies in amino acid sequence and tertiary structure of protein moleculesBiochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology, 1982
- The Cytosol of Human Erythrocytes Contains a Highly Ca2+-Sensitive Thiol Protease (Calpain I) and Its Specific Inhibitor Protein (Calpastatin)1The Journal of Biochemistry, 1981
- Isolation and Characterization of mRNA from Mammary Gland of Lactating CowAgricultural and Biological Chemistry, 1981
- Cloning DNA restriction endonuclease fragments with protruding single-stranded endsGene, 1980
- Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications.Proceedings of the National Academy of Sciences, 1979
- DNA sequencing with chain-terminating inhibitorsProceedings of the National Academy of Sciences, 1977