p36, the major cytoplasmic substrate of src tyrosine protein kinase, binds to its p11 regulatory subunit via a short amino-terminal amphiphatic helix.
Protein I is a hetero-tetramer which contains two copies each of p11 and p36. p36 (calpactin I, lipocortin II) is a major substrate of retrovirally encoded tyrosine protein kinases, while p11 modulates several Ca2+-induced properties also displayed by p36 alone. Here we have characterized the p11 binding site on p36 by fluorescence spectroscopy using porcine p36 labelled at cysteine 8 with the fluorophore Prodan (6-proprionyl-2-dimethylamino-naphthalene). We have used peptides of differing length from the amino-terminal domain of p36 to restrict the major binding site to the first 12 residues. Noticeable binding is still observed with a peptide containing only the first nine residues. Interestingly the N-terminal acetyl group of p36 forms a functional part of the p11 binding site. CD studies indicate that the binding region can form an alpha-helix, which seems to have amphiphatic properties when projected on a helical wheel. This structural element is also known for a calmodulin binding protein. Thus the question is raised whether other p11/calmodulin-related proteins interact with their target proteins via a similar mechanism. We also discuss how p11 could modulate p36 associated properties.
Johnsson
N
N
Max Planck Institute for Biophysical Chemistry, Göttingen, FRG.
Marriott
G
G
Weber
K
K
D000595:Amino Acid Sequence; D000818:Animals; D017302:Annexins; D001665:Binding Sites; D002135:Calcium-Binding Proteins / Q000032:analysis / Q000378:metabolism*; D002645:Chickens; D002850:Chromatography, Gel; D005454:Fluorescence Polarization; D006023:Glycoproteins / Q000032:analysis / Q000378:metabolism*; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D010455:Peptides / Q000138:chemical synthesis; D013050:Spectrometry, Fluorescence; D013552:Swine; D016704:Synapsins
D017302:Annexins; D002135:Calcium-Binding Proteins; D006023:Glycoproteins; D009419:Nerve Tissue Proteins; D010455:Peptides; D016704:Synapsins