Evidence for multisite ADP-ribosylation of neuronal phosphoprotein B-50/GAP-43.
The neuronal phosphoprotein B-50/GAP-43 is associated with neuronal growth and regeneration and is involved in the calcium/CaM and G(o) signal transduction systems. In particular, B-50 interacts uniquely with CaM by binding in the absence of Ca2+. Previously identified as a major neuronal substrate for protein kinase C, which releases CaM via phosphorylation, B-50 has more recently been shown to be a substrate for endogenous ADP-ribosyltransferases. In the present study, we utilized amino acid modification with iodoacetamide and chemical stability to mercury and neutral hydroxylamine to demonstrate that the predominant site of ADP-ribosylation is Cys 3 and/or Cys 4. Chymotryptic peptide mapping further revealed a second, less labelled site of ribosylation in the C-terminal region. The results also demonstrate that, in contrast to PKC phosphorylation, ADP-ribosylation of B-50 does not mediate CaM binding. Since Cys 3 and Cys 4, by palmitoylation, are important for membrane anchoring, our findings suggest that ADP-ribosylation of B-50 may have a role in directing the intracellular localization of the protein. Hence, ribosylation of B-50 may mediate where B-50 interacts with signal transduction pathways.
Philibert
K
K
Department of Medical Physiology, University of Calgary, Health Sciences Centre, Alberta, Canada.
Zwiers
H
H
D000246:Adenosine Diphosphate Ribose / Q000378:metabolism*; D000818:Animals; D002147:Calmodulin / Q000378:metabolism; D002148:Calmodulin-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D003545:Cysteine / Q000737:chemistry; D019922:GAP-43 Protein; D007460:Iodoacetamide / Q000737:chemistry; D008297:Male; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley
D002147:Calmodulin; D002148:Calmodulin-Binding Proteins; D019922:GAP-43 Protein; D008562:Membrane Glycoproteins; D009419:Nerve Tissue Proteins; D000246:Adenosine Diphosphate Ribose; D003545:Cysteine; D007460:Iodoacetamide