Amino acid sequences and Ca2(+)-binding properties of two isoforms of barnacle troponin C.
Much of our knowledge of the relationship between elevation of free sarcoplasmic [Ca2+] and skeletal muscle contraction has come from physiological studies on barnacle muscle fibers. Little is known, however, about the biochemical properties of the barnacle proteins responsible for Ca2+ regulation. In order to help rectify this unfortunate situation, we purified the two major isoforms (BTnC1 and BTnC2) of troponin C (TnC) from the giant barnacle, Balanus nubilis, and determined their amino acid sequences. BTnC2, the more abundant isoform, contains 151 amino acid residues and has a calculated molecular weight of 16,838. Due to an elongated N-terminus, BTnC1 contains 158 amino acid residues and has a calculated molecular weight of 17,984. The two sequences can be aligned from their C-termini, with no insertions or deletions, but they are only 61% identical. Sequence differences are twice as frequent in the N-terminal halves as in the C-terminal halves but occur in all regions of the polypeptides. This indicates that BTnC1 and BTnC2 are products of different genes, rather than alternative transcripts of a single gene. Both isoforms contain the usual four ancestral Ca2(+)-binding regions, numbered I-IV from the N-terminus. Analysis of the sequences predicts that functional Ca2(+)-binding sites are present only in regions II and IV and that these sites are low-affinity Ca2(+)-specific type sites. Direct Ca2(+)-binding measurements using fluorescent Ca2+ indicators show that both isoforms bind 2 Ca2+/mol with equal affinity (KCa = 1.3 x 10(5) M-1) and the sites appear to be Ca2(+)-specific.
Collins
J H
JH
Department of Biological Chemistry, School of Medicine, University of Maryland, Baltimore 21201.
Theibert
J L
JL
Francois
J M
JM
Ashley
C C
CC
Potter
J D
JD
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism*; D002645:Chickens; D007536:Isomerism; D008969:Molecular Sequence Data; D009119:Muscle Contraction; D001468:Thoracica / Q000378:metabolism*; D014336:Troponin / Q000737:chemistry / Q000378:metabolism*; D019209:Troponin C
D014336:Troponin; D019209:Troponin C; D002118:Calcium