title: Alkylation of cysteine 82 of p11 abolishes the complex formation with the tyrosine-protein kinase substrate p36 (annexin 2, calpactin 1, lipocortin 2).
abstract: p36 (annexin 2) is the major cytoplasmic target of the src tyrosine-kinase and forms in vitro and in vivo a stable tetrameric complex in which two p36 polypeptides interact with a dimer of a unique p11 polypeptide. p11 belongs into the superfamily of EF-hand proteins. Upon mild cysteine modification conditions, both cysteines (position 61 and 82) of the free p11 become substituted, and the ability to form the p36.p11 complex is lost. Under the same conditions, the 2 cysteines of p11 incorporated into the complex display differential reactivity. Here, cysteine 61 is fully substituted while cysteine-82 is protected. p11 derivatives substituted only on cysteine 61 retain binding activity for p36 unless cysteine 82 is substituted by a second cycle of modification of the isolated p11. Thus, the C-terminal extension protruding from the second EF-hand of the p11 molecule (residues 77-96) is important for the interaction with p36. As a consequence of our analysis, we report a new separation of p36 and p11 from the p36.p11 complex. This is based on a reversible cysteine modification and thus is an alternative to the denaturation and renaturation cycle used previously.
authors: Max Planck Institute for Biophysical Chemistry, Göttingen, Federal Republic of Germany.
authors: N
authors: N
authors: Johnsson
authors: K
authors: K
authors: Weber
mesh_terms: D000478:Alkylation; D000818:Animals; D017302:Annexins; D001665:Binding Sites; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D003545:Cysteine; D006023:Glycoproteins / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D011485:Protein Binding; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D013379:Substrate Specificity; D013552:Swine
chemical_list: D017302:Annexins; D002135:Calcium-Binding Proteins; D006023:Glycoproteins; D008565:Membrane Proteins; D011505:Protein-Tyrosine Kinases; D003545:Cysteine