Ca2+-dependent interaction with calmodulin is conserved in the synapsin family: identification of a high-affinity site.
The synapsins are a family of proteins associated with small synaptic vesicles that are implicated in synaptic maintenance and in the supply of vesicles for exocytosis. They are well characterized as substrates for protein kinases, and one class of synapsin, synapsin I, has been shown to bind, and be regulated by, calmodulin. A representative of the synapsin II class is now shown to bind calmodulin. Optical biosensor assays of Ca2+-dependent calmodulin binding to recombinant rat synapsin IIb indicated an apparent KD for calmodulin of 31 +/- 5 nM. Phosphorylation at Ser 10 increased the rates of calmodulin association (by a factor of 10) and dissociation (by a factor of 20). Fragment analysis and predictions from the sequence indicated two potential calmodulin binding sequences in the conserved central (C) domain. Peptides representing these sequences (residues 122-143 and 313-334 in synapsin IIb) were synthesized. Peptide 122-143 was found to bind calmodulin (KD 32 +/- 10 nM) and inhibit interaction of synapsin IIb with calmodulin. The interaction of peptide 313-334 was much weaker. Sequences similar to residues 122-143 are present in all published synapsin sequences. Calmodulin binding by synapsins seems not to be confined to mammals: a recombinant Drosophila synapsin 1 fragment containing part of the C-domain showed Ca2+-dependent binding to mammalian calmodulin. We conclude that calmodulin binding to synapsins is likely to be a general aspect of regulation of synaptic function.
Nicol
S
S
Research School of Biosciences, University of Kent, Canterbury, Kent, CT2 7NJ, England.
Rahman
D
D
Baines
A J
AJ
D006631:Amine Oxidase (Copper-Containing) / Q000235:genetics / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D015374:Biosensing Techniques; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism; D011485:Protein Binding; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D013045:Species Specificity; D016704:Synapsins / Q000145:classification / Q000235:genetics / Q000378:metabolism*
D002147:Calmodulin; D010446:Peptide Fragments; D011994:Recombinant Proteins; D016704:Synapsins; D006631:Amine Oxidase (Copper-Containing); D002118:Calcium