Interaction of tubulin with drugs and alkylating agents. 1. Alkylation of tubulin by iodo[14C]acetamide and N,N'-ethylenebis(iodoacetamide).
The sulfhydryl groups of tubulin are reported to play a role in regulating microtubule assembly and colchicine binding to tubulin. The alkylating agents iodo[14C]acetamide and its bifunctional analogue N,N'-ethylenebis(iodoacetamide) are used as probes for the sulfhydryl groups of tubulin. In the presence of 8 M urea, alpha- and beta-tubulin have 10-11 and 8 alkylatable sulfhydryls, respectively, and one of the high molecular weight proteins (HMW 2) has 5 sulfhydryls/Mr 271 000. In the absence of urea, the rates of alkylation of alpha- and beta-tubulin are significantly lower but that of HMW 2 is unaffected. The sulfhydryls of tubulin are masked in intact microtubules. N,N'-Ethylenebis(iodoacetamide) reacts with free tubulin to generate a band, designated beta, which migrates ahead of beta on polyacrylamide gels. beta appears to represent a form of beta-tubulin containing at least one intrachain cross-link between sulfhydryl groups. Formation of beta* is inhibited in intact microtubules and is abolished if tubulin is denatured by 8 M urea, 1% sodium dodecyl sulfate, or boiling. N,N'-Ethylenebis(iodoacetamide) may thus be used as a probe for the native conformation of free tubulin.
D000477:Alkylating Agents; D000818:Animals; D001921:Brain / Q000378:metabolism; D002250:Carbon Radioisotopes; D002417:Cattle; D003432:Cross-Linking Reagents; D005029:Ethylenediamines; D007460:Iodoacetamide / Q000031:analogs & derivatives; D007461:Iodoacetates; D007700:Kinetics; D008870:Microtubules / Q000378:metabolism; D010446:Peptide Fragments / Q000032:analysis; D011485:Protein Binding; D014404:Tubulin / Q000378:metabolism
D000477:Alkylating Agents; D002250:Carbon Radioisotopes; D003432:Cross-Linking Reagents; D005029:Ethylenediamines; D007461:Iodoacetates; D010446:Peptide Fragments; D014404:Tubulin; C032156:N,N'-ethylenebis(iodoacetamide); D007460:Iodoacetamide