The binding sites of rabbit skeletal troponin-I on troponin-T

1980 ◽  
Vol 58 (8) ◽  
pp. 649-654 ◽  
Author(s):  
Joyce R. Pearlstone ◽  
Lawrence B. Smillie
Keyword(s):  
Skeletal Muscle ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Binding Site ◽  
Binding Sites ◽  
Troponin I ◽  
Troponin T ◽  
Gel Filtration ◽  
Troponin C ◽  
Separate Site

Various fragments derived from rabbit skeletal muscle troponin-T (Tn-T) by chemical and (or) proteolytic cleavage were mixed with whole troponin-I (Tn-I) and applied to a Sephadex G-75 gel filtration column in order to determine the binding site of Tn-I on Tn-T. This site of interaction was found to span two distinct regions of Tn-T. The first site involves the highly acidic NH2-terminal fragment CB3 (residues 1–70 of Tn-T). A second separate site is located in the region of residues 152–209 of Tn-T. The present study, in conjunction with our earlier work on tropomyosin – Tn-T binding and Tn-T – troponin-C binding, depicts Tn-T as being a functionally efficient molecule composed of several distinct domains of specialized amino acid sequence, each of which carries out a role in the binding of a different protein.

scholarly journals Biological activities of bovine cardiac-muscle troponin C C-terminal peptide (residues 84-161)

1982 ◽  
Vol 207 (2) ◽  
pp. 185-192 ◽  
Author(s):  
N V Barskaya ◽  
N B Gusev
Keyword(s):  
Skeletal Muscle ◽  
Amino Acid ◽  
Cardiac Muscle ◽  
Sodium Dodecyl Sulphate ◽  
Binding Sites ◽  
Binding Constant ◽  
Troponin I ◽  
Biological Activities ◽  
Sodium Dodecyl ◽  
Troponin C

1. Bovine cardiac-muscle troponin C was digested at cysteine residues 35 and 84, and the C-terminal peptide (residues 84-161) was isolated. 2. The C-terminal peptide contains two Ca2+-binding sites. These sites bind Ca2+ with a binding constant of 2.0×10(8) M-1. In the presence of 2 mM-Mg2+ the binding constant for Ca2+ is decreased to 3.70×10(7) M-1. The corresponding constants for native troponin C are 5.90×10(7) M-1. and 2.90×10(7) M-1 respectively. 3. Electrophoretic mobility of the C-terminal peptide is increased in the presence of 0.1 mM-CaCl2 as compared with the mobility in the presence of 2mM-EDTA. The same phenomenon was observed when electrophoresis was performed in the presence of 6 M-urea or 0.1% sodium dodecyl sulphate. 4. When saturated with Ca2+, the C-terminal peptide forms complexes with bovine cardiac-muscle troponin I both in the absence and in the presence of 6 M-urea. This complex is dissociated on removal of Ca2+. 5. The data suggest that the C-terminal peptide of troponin C contains two Ca2+/Mg2+-binding sites and interacts with troponin I. Thus, despite the 30% difference in amino acid composition, the properties of bovine cardiac-muscle troponin C C-terminal peptide are similar to those of rabbit skeletal-muscle troponin C C-terminal peptide.

scholarly journals The primary structure of troponin T and the interaction with tropomyosin

1975 ◽  
Vol 151 (1) ◽  
pp. 85-97 ◽  
Author(s):  
P Jackson ◽  
G W Amphlett ◽  
S V Perry
Keyword(s):  
Skeletal Muscle ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Primary Structure ◽  
Troponin T ◽  
Troponin C ◽  
The Other ◽  
Tryptic Digestion ◽  
Partial Digestion ◽  
Common Sequence

1. Eight peptides were separated from the CNBr digest of troponin T from rabbit white skeletal muscle and characterized. 2. By study of the amino acid sequence of the methionine-containing peptides isolated after chymotryptic and tryptic digestion and of the N- and C-terminals of the CNBr peptides, six of the latter were shown to be arranged in the sequence CNB1-CNB2-CNB5-CNB6-CNB8-CNB7. The other two peptides, CNB1′ and CNB3, have been shown to be partial digestion products. 3. The CNBr peptides CNB1′ and CNB2 contained a common sequence and were the only peptides in CNBr digests of troponin T that formed a complex with tropomyosin as judged by viscometric and electrophoretic studies. 4. It is concluded that tropomyosin interacts with the N-terminal half of the troponin T molecule approximately in the region lying between residues 70 and 160. 5. Electrophoretic evidence indicates that tropomyosin and troponin C interact with troponin T. 6. None of the major CNBr peptides of troponin T isolated formed a complex with troponin C on electrophoresis at pH 8.6.

scholarly journals Proteolytic fragments of troponin C. Localization of high and low affinity Ca2+ binding sites and interactions with troponin I and troponin T

1978 ◽  
Vol 253 (15) ◽  
pp. 5452-5459
Author(s):  
P.C. Leavis ◽  
S.S. Rosenfeld ◽  
J. Gergely ◽  
Z. Grabarek ◽  
W. Drabikowski
Keyword(s):  
Binding Sites ◽  
Troponin I ◽  
Troponin T ◽  
Troponin C

The interaction of rabbit skeletal muscle troponin-T fragments with troponin-I

1985 ◽  
Vol 63 (3) ◽  
pp. 212-218 ◽  
Author(s):  
Joyce R. Pearlstone ◽  
Lawrence B. Smillie
Keyword(s):  
Skeletal Muscle ◽  
Ionic Strength ◽  
Cyanogen Bromide ◽  
Troponin I ◽  
Polypeptide Chain ◽  
Troponin T ◽  
Gel Filtration ◽  
Equimolar Amount ◽  
Binding Region ◽  
Helical Content

The interactions of troponin-I (Tn-I) with a variety of fragments spanning the length of the troponin-T (Tn-T) polypeptide chain have been reinvestigated at physiological ionic strength by affinity chromatographic, gel filtration, and circular dichroism methodologies. Strong binding was observed with fragment T2 (residues 159–259) mimicking that observed with whole Tn-T and Tn-I. Partial binding was seen with the shorter cyanogen bromide (CB) fragments of Tn-T in the order CB4 (residues 176–230) > CB6 (residues 239–259) or CB5 (residues 152–175). No interaction with Tn-I was observed with fragments (CB2, CB3, T1) encompassing residues 1–158 of Tn-T. Based on the present results and the work of others, the binding region for Tn-I includes residues 159–259 and perhaps extends into the highly helical CB2 region (residues 71–151) of Tn-T. No evidence has been obtained by ourselves or others for the interaction of the CB3 region (1–70) with Tn-I. A significant increase (11.6%) in α-helical content was observed when an equimolar amount of fragment T2 (residues 159–259) was mixed with Tn-I, a result similar to that seen with whole Tn-T and Tn-I.

scholarly journals The amino acid sequence of rabbit cardiac troponin I

1976 ◽  
Vol 159 (3) ◽  
pp. 633-641 ◽  
Author(s):  
R J A. Grand ◽  
J M Wilkinson ◽  
L E More
Keyword(s):  
Skeletal Muscle ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Inhibitory Activity ◽  
Cardiac Troponin ◽  
Troponin I ◽  
Muscle Protein ◽  
Cardiac Troponin I ◽  
Skeletal Muscle Protein ◽  
Fast Skeletal Muscle

The complete amino acid sequence of troponin I from rabbit cardiac muscle was determined by the isolation of four unique CNBr fragments, together with overlapping tryptic peptides containing radioactive methionine residues. Overlap data for residues 35-36, 93-94 and 140-145 are incomplete, the sequence at these positions being based on homology with the sequence of the fast-skeletal-muscle protein. Cardiac troponin I is a single polypeptide chain of 206 residues with mol.wt. 23550 and an extinction coefficient, E 1%,1cm/280, of 4.37. The protein has a net positive charge of 14 and is thus somewhat more basic than troponin I from fast-skeletal muscle. Comparison of the sequences of troponin I from cardiac and fast skeletal muscle show that the cardiac protein has 26 extra residues at the N-terminus which account for the larger size of the protein. In the remainder of sequence there is a considerable degree of homology, this being greater in the C-terminal two-thirds of the molecule. The region in the cardiac protein corresponding to the peptide with inhibitory activity from the fast-skeletal-muscle protein is very similar and it seems unlikely that this is the cause of the difference in inhibitory activity between the two proteins. The region responsible for binding troponin C, however, possesses a lower degree of homology. Detailed evidence on which the sequence is based has been deposited as Supplementary Publication SUP 50072 (20 pages), at the British Library Lending Division, Boston Spa, Wetherby, West Yorkshire LS23 7QB, U.K., from whom copies may be obtained on the terms given in Biochem. J. (1976) 153, 5.

scholarly journals The amino acid sequence of troponin I from rabbit skeletal muscle

1975 ◽  
Vol 149 (2) ◽  
pp. 493-496 ◽  
Author(s):  
J M Wilkinson ◽  
R J A. Grand
Keyword(s):  
Skeletal Muscle ◽  
Molecular Weight ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Cyanogen Bromide ◽  
Troponin I ◽  
Rabbit Skeletal Muscle ◽  
British Library ◽  
Amino Acid Residues ◽  
N Terminus

The complete amino acid sequence of rabbit skeletal muscle troponin I was determined by the isolation of the cyanogen bromide fragments and the tryptic methionine-containing peptides. Troponin I contains 179 amino acid residues and has a molecular weight of 20864. Its N-terminus is acetylated. Detailed evidence on which the sequence is based has been deposited as Supplementary Publication SUP 50055 (23 pages) at the British Library (Lending Division), Boston Spa, Wetherby, West Yorkshire LS23 7QB, U.K., from whom copies may be obtained on the terms given in Biochem. J. (1975) 145, 5.

Sign in / Sign up

Export Citation Format

Share Document