Mirror image motifs mediate the interaction of the COOH terminus of multiple synaptotagmins with the neurexins and calmodulin.
I have previously reported that the COOH-terminal 34 amino acids of synaptotagmin 1 are capable of interacting with the presynaptic proteins, the neurexins. Multiple synaptotagmins and a synaptotagmin-like protein, rabphilin 3A, are conserved in this domain, raising the possibility that many different synaptotagmins may interact with neurexins. Here 1 report that the COOH termini of synaptotagmins 1, 2, 4, 5, 6, 7, and 9 and rabphilin 3A are capable of interacting with neurexins. The COOH terminus of rabphilin 3A is still capable or substantial enrichment of neurexins from solubilized brain membranes even though only 11 of 33 residues are identical with the COOH terminus of synaptotagmin 1. Like the purification of neurexins on the COOH terminus of synaptotagmin 1, purification by the COOH terminus of rabphilin 3A is calcium-independent. The conservation between carboxyl termini of these proteins suggests symmetrical motifs are necessary for neurexin binding. These include the sequence Leu-X-His-Trp, followed by 13 amino acids, and the sequence Trp-His-X-Lcu. Deletion of the first motif or substitution of residues in the second of these motifs greatly reduces neurexin enrichment. Interestingly, these same COOH termini yield substantial calcium-dependent enrichment of calmodulin mediated by the first of these sequence motifs. This correlates with the binding of 125I-labeled calmodulin by recombinant pieces of synaptotagmn 1 containing the carboxyl terminus. These data suggest that multiple synaptotagmins may interact with neurexins to mediate docking or regulation of neurotransmitter release and that synaptotagmins may be calcium-regulated via interaction with calmodulin.
Division of Neuroscience, Baylor College of Medicine, Houston, Texas 77030, USA.
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D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D001921:Brain / Q000378:metabolism; D002135:Calcium-Binding Proteins; D002147:Calmodulin / Q000737:chemistry / Q000378:metabolism; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D017124:Conserved Sequence / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D019204:GTP-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000378:metabolism; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D016415:Sequence Alignment; D013572:Synaptic Vesicles / Q000378:metabolism; D050863:Synaptotagmin I; D050857:Synaptotagmins; D033921:Vesicular Transport Proteins; D020691:rab GTP-Binding Proteins; D000097624:Rabphilin-3A
D048868:Adaptor Proteins, Signal Transducing; D002135:Calcium-Binding Proteins; D002147:Calmodulin; D019204:GTP-Binding Proteins; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D010446:Peptide Fragments; D011994:Recombinant Proteins; D050863:Synaptotagmin I; D050857:Synaptotagmins; D033921:Vesicular Transport Proteins; D020691:rab GTP-Binding Proteins; D000097624:Rabphilin-3A; C494312:Syt1 protein, rat