Chemical modification of bovine brain tubulin with the guanine nucleotide affinity analogue 5'-p-fluorosulfonylbenzoylguanosine.
Bovine brain tubulin purified in the absence of GTP and MgCl2, reacts with 5'-p-fluorosulfonylbenzoylguanosine (5'-FSBG)2, an affinity analog of GTP and two moles of the reagent are incorporated per mole of tubulin at 0 degree C. 5'-FSBG is unable to promote the polymerization of tubulin into microtubules. 2 mM GTP, podophyllotoxin and vinblastine provide almost 50% protection against the modification, when added individually. Combination of these ligands gives maximal protection. Tubulin modified with 5'-FSBG lost two sulfhydryl groups per mole of tubulin and reduction with beta-mercaptoethanol led to the loss of the 2 moles of FSBG that had been incorporated. These data are interpreted on the basis that the modification of tubulin by 5'-FSBG proceeds via a thiosulfonate intermediate between the analogue and a reactive thiol group at or near that portion of the GTP binding site of tubulin where the phosphate moiety of GTP binds.
D000345:Affinity Labels / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001923:Brain Chemistry; D002417:Cattle; D006151:Guanosine / Q000031:analogs & derivatives* / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000494:pharmacology; D008870:Microtubules / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000032:analysis; D014404:Tubulin / Q000378:metabolism*
C017320:5'-(4-fluorosulfonylbenzoyl)guanosine; D000345:Affinity Labels; D013438:Sulfhydryl Compounds; D014404:Tubulin; D006151:Guanosine; D006160:Guanosine Triphosphate