Detergents and peptides alter proteolysis and calmodulin binding of B-50/GAP-43 in vitro.
The neuronal growth-associated protein B-50/GAP-43 is a substrate for protein kinase C, binds to calmodulin in a calcium-independent manner, and in vitro is subject to an endogenous and chymotrypsin-mediated hydrolysis in the vicinity of the single kinase C phosphorylation site. All of these processes can be influenced by corticotrophin (ACTH). In the present study we have investigated whether these biochemical interactions involving B-50 could have common structural determinants. Chymotryptic digestion of B-50 in the presence or absence of a nonionic detergent and ACTH demonstrated that hydrolysis is potentiated by a lipid-like environment that primarily affects the protein rather than the protease or the peptide. Furthermore, this lipid dependency appears to extend to the binding of dephosphorylated B-50 to calmodulin, which appears to occur only in the presence of a nonionic detergent or lipid and the absence of calcium. A structure-activity study for ACTH-mediated inhibition of B-50 proteolysis by an endogenous protease that copurifies with B-50 in a detergent extract of synaptosomal plasma membranes showed that ACTH1-24, ACTH5-24, ACTH5-16, dynorphin, and corticostatin inhibited the conversion of rat B-50 to B-5041-226. In contrast, ACTH7-16, Org2766, and neurotensin had no detectable effect on B-50 proteolysis at concentrations of 10 and 50 microM.(ABSTRACT TRUNCATED AT 250 WORDS)
Coggins
P J
PJ
Department of Medical Physiology, University of Calgary, Alberta, Canada.
Zwiers
H
H
D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000378:metabolism*; D002147:Calmodulin / Q000302:isolation & purification / Q000378:metabolism*; D002417:Cattle; D002918:Chymotrypsin / Q000494:pharmacology*; D019922:GAP-43 Protein; D008297:Male; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D016900:Neurofilament Proteins / Q000378:metabolism*; D010455:Peptides / Q000494:pharmacology*; D010766:Phosphorylation; D011092:Polyethylene Glycols / Q000494:pharmacology*; D011485:Protein Binding; D011493:Protein Kinase C / Q000302:isolation & purification / Q000378:metabolism; D017433:Protein Structure, Secondary; D051381:Rats; D017207:Rats, Sprague-Dawley; D013570:Synaptic Membranes / Q000378:metabolism; D013574:Synaptosomes / Q000378:metabolism*
D002147:Calmodulin; D019922:GAP-43 Protein; D008562:Membrane Glycoproteins; D009419:Nerve Tissue Proteins; D016900:Neurofilament Proteins; D010455:Peptides; D011092:Polyethylene Glycols; C005092:lubrol; D011493:Protein Kinase C; D002918:Chymotrypsin