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J G Demaille

Publications and source records attributed to J G Demaille.

At least 73 records · Page 4Linked to original sources
PubMed1980

Homologous calcium-binding proteins in the activation of skeletal, cardiac, and smooth muscle myosin light chain kinases.

In order to identify the physiological regulator of calcium dependent myosin light chain kinases of cardiac, skeletal, and smooth muscles, the effects of the three homologous calciproteins, calmodulin, troponin C, and parvalbumin, on the kinases isolated from bovine myocardium, rabbit skeletal muscle, and turkey gizzard were examined. Only calmodulin was effective in stimulating the cardiac, skeletal, or smooth muscle kinase; troponin C and parvalbumin exhibited no activation of any of the three kinases, even when examined at concentrations as high as 10-(5) M. It is concluded that calmodulin is the specific regulator of myosin light chain kinase in cardiac, skeletal, and smooth muscle.

Adenosine Triphosphatases↗
PubMed1979

Purification and characterization of bovine cardiac calmodulin-dependent myosin light chain kinase.

Myosin light chain kinase, which is located primarily in the soluble fraction of bovine myocardium, has been isolated and purified approximately 1200-fold with 16% yield by a three-step procedure. The approximate content of soluble myosin light chain kinase in heart is calculated to be 0.63 microM. The isolated kinase is active only as a ternary complex consisting of the kinase, calmodulin, and Ca2+; the apparent Kd for calmodulin is 1.3 nM. The enzyme also exhibits a requirement for Mg2+ ions. Myosin light chain kinase is a monomeric enzyme with Mr = 85,000. The enzyme exhibits a Km for ATP of 175 microM, and a K0.5 for the regulatory light chain of cardiac myosin of 21 microM. The optimum pH is 8.1. Kinase activity is specific for the regulatory light chain of myosin. The specific activity of the isolated enzyme (30 nmol 32P/min/mg of protein) is considerably less than and corresponding values reported for the skeletal and smooth muscle light chain kinases. This is probably due to proteolysis during extraction of the myocardium, a phenomenon which has, as yet, proven impossible to eliminate. In contrast to the smooth muscle enzyme (Adelstein, R.S., Conti, M.A., Hathaway, D.R., and Klee, C.B. (1978) J. Biol. Chem. 253, 8347-8350), the cardiac kinase is not phosphorylated by the catalytic subunit of cAMP-dependent protein kinase.

Animals↗
PubMed1979

Sequence of two phosphorylated sites in the catalytic subunit of bovine cardiac muscle adenosine 3':5'-monophosphate-dependent protein kinase.

The sequences of two phosphopeptides isolated from the catalytic subunit of bovine cardiac muscle cAMP-dependent protein kinase (type II) and from two of its cyanogen bromide fragments, have been determined. One phosphorylation site is a threonyl residue located approximately 180 residues from the blocked NH2 terminus. Its sequence is: -Gly-Arg-Thr-Trp-Thr(P)-Leu-Cys- and includes one of the three sulfhydryl groups present in the molecule. The second phosphorylated site within the sequence: -Val-Ser(P)-Ile-Asn- is located towards the carboxyl end of the protein where the other 2 cysteinyl residues also reside. The finding that phosphorylation of the catalytic subunit occurs on two discrete sites rather than at random suggests that it might be of physiological importance, e.g. in the regulation of enzyme activity.

Amino Acid Sequence↗
PubMed1979

Evolutionary diversification of structure and function in the family of intracellular calcium-binding proteins.

The maximum parsimony method was used to reconstruct the genealogical history of the family of intracellular calcium-binding proteins represented by six major present-day lineages, three of which--calcium dependent modulator protein, heart and skeletal muscle troponin Cs, and alkali light chains of myosin--were found to share a closer kinship with one another than with the other lineages. Similarly, parvalbumins and regulatory light chains of myosin were depicted as more closely related, whereas the branch of intestinal calcium-binding protein proved to have the most distant separation. The computer-generated amino acid sequence for the common ancestor of these six lineages described a four domain protein in which each domain of approximately 40 amino acid residues had a mid-region. 12 residue segment that bound calcium and had properties most resembling those of the calcium dependent modulator protein. It could then be deduced that parvalbumins evolved by deletion of domain I, inactivation of calcium-binding properties in domain II, and acquisition of increased affinity for Ca++ and Mg++ in domains III and IV. Regulatory light chains of myosin lost the cation binding property from three domains, retaining it in I, whereas alkali light chains of myosin lost this ability from each of the four domains. In skeletal muscle troponin C all domains retained their calcium-binding activity; however, like parvalbumins, domains III and IV acquired high affinity properties. Cardiac troponin C lost its binding activity from domain I but otherwise resembled the skeletal muscle form. Finally, intestinal calcium-binding protein evolved by deletion of domains III and IV. Positive selection could be implicated in these evolutionary changes in that the rate of fixation of mutations substantially increased in the mid portions of those domains which were loosing calcium-binding activity. Likewise, when the cation binding sites were changing from low to high affinity, an accelerated rate of fixed mutations was observed. Once this new functional parameter was selected these regions showed a remarkable conservatism, as did those binding sites which were maintaining the lower affinity. Moreover even in sequence regions not directly involved in cation binding, the lineage of troponin C because very conservative over the past 300 million years, perhaps becuase of the necessity for maintaining specific interfaces in order for the molecule to interact with troponin I and T in a functional thin myofilament. A similar phenomenon was observed in domain II of the regulatory light chains of the myosin lineage suggesting a possible binding site with the heavy chain of myosin.

Amino Acid Sequence↗
PubMed1979

The protein inhibitor of adenosine 3':5'-monophosphate-dependent protein kinases. Isolation and characterization of three isoinhibitors.

The protein inhibitor of adenosine 3':5 monophosphate-dependent protein-kinases has been purified from rabbit skeletal muscle by affinity chromatography on Sepharose-bound catalytic subunit of the kinase (Demaille et al. (1977) Biochemistry 16, 3080-3086). The inhibitory material can be separated into three species A, B and C either by polyacrylamide gel electrophoresis or by anion-exchange on DEAE-cellulose. However, the isoinhibitors display the same molecular weight and isoelectric point, and the same absence of acid-stable covalently bound phosphate. Their amino acid compositions are strikingly similar and their NH2-terminus are blocked. Also, their COOH-terminus appear to be very close to one another when studied by carboxypeptidase Y digestion. Their major difference lies in their inhibitory properties which are significantly weaker in inhibitor C (Ki = 0.87 nM) than in A and B (Ki = 0.18 and 0.23 nM, respectively). This is the first report on the existence of various forms of the protein-kinase inhibitor that exhibit different inhibitory properties and may play a role in the regulation of the protein-kinase system.

Amino Acid Sequence↗
PubMed1979

Calcium and magnesium binding by parvalbumin. A proton magnetic resonance spectral study.

Proton magnetic resonance spectroscopy has been used to monitor the kinetics and nature of the conformational transitions induced by binding of calcium and magnesium to carp parvalbumin. Rate constants have been determined for the exchange between the cation dependent conformational states of the protein in solution. These data enable a description of the kinetics and mechanism controlling the calcium flux in vivo during contraction.

Animals↗