A calmodulin-binding sequence in the C-terminus of human cardiac titin kinase.
The giant muscle proteins of the titin family, which are specific for the striated muscles of vertebrates and invertebrates, contain as a common feature a catalytic protein kinase domain of so far unclear function and regulation. In myosin light chain kinase, a family evolutionarily related to titin, kinase regulation is achieved by calmodulin binding to a region of the kinase C-terminus which bears similarity to the substrate. A calmodulin-binding sequence has also been identified in the C-terminus of the Aplysia twitchin kinase. In analogy, we identified a putative calmodulin-binding site in the titin kinase C-terminal sequence. The expressed catalytic domain itself and a series of synthetic peptides from this region were tested for their ability to bind calmodulin. Biochemical data indicate that titin kinase as well as peptides from its C-terminus bind to calmodulin in an equimolar complex in the presence of calcium. The interaction of truncated peptides with calmodulin is, however, weaker than that of myosin light chain kinase. Nuclear magnetic resonance studies showed that these peptides have a tendency to adopt alpha-helical conformations in solution. Helicity increases upon binding of calmodulin in a calcium-dependent fashion, as judged by circular dichroism spectra. We, therefore, propose that this calmodulin-binding region of titin could play a regulatory role for the enzyme, the substrate of which still remains to be identified.
Gautel
M
M
EMBL, Heidelberg, Germany.
Castiglione Morelli
M A
MA
Pfuhl
M
M
Motta
A
A
Pastore
A
A
D000595:Amino Acid Sequence; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism*; D002148:Calmodulin-Binding Proteins / Q000378:metabolism*; D002942:Circular Dichroism; D064211:Connectin; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000737:chemistry / Q000378:metabolism*; D009206:Myocardium / Q000201:enzymology*; D011485:Protein Binding; D011487:Protein Conformation; D011494:Protein Kinases / Q000737:chemistry / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid
D002147:Calmodulin; D002148:Calmodulin-Binding Proteins; D064211:Connectin; D009124:Muscle Proteins; C577285:TTN protein, human; D011494:Protein Kinases; D002118:Calcium