Different effects of tubulin ligands on the intrachain cross-linking of beta 1-tubulin.
When bovine brain tubulin purified in the absence of GTP and MgCl2 is reacted with N,N'-ethylene-bis(iodoacetamide) (EBI), a bifunctional analogue of iodoacetamide, three new electrophoretically distinct species of tubulin are generated, migrating ahead of beta 1-tubulin on gels containing Na dodecyl sulfate. All three bands appear to be derived from the beta 1 subunit of tubulin and not from the alpha or beta 2 subunit. Accordingly, the bands have been designated beta 1 s, beta 1, and beta 1s in order of increasing electrophoretic mobility. EBI appears to introduce two intrachain cross-links into beta 1-tubulin; the beta 1s band contains one of these cross-links, designated beta s, the beta 1 band contains the other cross-link, designated beta s, and the beta 1 s band contains both cross-links. Both cross-links appear to involve sulfhydryl groups. Colchicine, podophyllotoxin, and nocodazole completely inhibit beta formation while GTP, vinblastine, and maytansine enhance it. It contrast, formation of beta s is completely blocked by guanine nucleotides and by maytansine, while vinblastine inhibits this by 70%. Colchicine, podophyllotoxin, and nocodazole enhance beta s formation. These results show that tubulin has the unusual property of having two discrete sites which can be targeted by an alkylating agent with each site having its alkylation inhibited by a different set of ligands. The results are consistent with several models, including one where vinblastine and maytansine have overlapping binding sites on the beta-subunit of tubulin relatively close to the GTP binding site.
D000818:Animals; D001562:Benzimidazoles / Q000494:pharmacology; D001923:Brain Chemistry; D002417:Cattle; D003078:Colchicine / Q000494:pharmacology; D003432:Cross-Linking Reagents / Q000494:pharmacology*; D005029:Ethylenediamines / Q000494:pharmacology*; D005260:Female; D006160:Guanosine Triphosphate / Q000494:pharmacology; D008024:Ligands / Q000494:pharmacology; D008274:Magnesium / Q000494:pharmacology; D015636:Magnesium Chloride; D008453:Maytansine / Q000494:pharmacology; D015739:Nocodazole; D011034:Podophyllotoxin / Q000494:pharmacology; D014404:Tubulin / Q000378:metabolism; D014747:Vinblastine / Q000494:pharmacology
D001562:Benzimidazoles; D003432:Cross-Linking Reagents; D005029:Ethylenediamines; D008024:Ligands; D014404:Tubulin; D015636:Magnesium Chloride; D008453:Maytansine; D014747:Vinblastine; C032156:N,N'-ethylenebis(iodoacetamide); D006160:Guanosine Triphosphate; D008274:Magnesium; D011034:Podophyllotoxin; D015739:Nocodazole; D003078:Colchicine