Assembly and colchicine binding characteristics of tubulin with maximally tyrosinated and detyrosinated alpha-tubulins.
The posttranslational removal and readdition of tyrosine at the C-terminus of alpha-tubulin is associated with generation of microtubule populations that differ in intracellular distributions, turnover rates, and sensitivities to microtubule-depolymerization agents. Here, we compared the in vitro assembly and colchicine binding characteristics of tubulin dimer preparations composed of alpha-tubulin that had been maximally tyrosinated (approximately 40% tyrosinated) by tubulin-tyrosine ligase and maximally detyrosinated (100% detyrosinated) by carboxypeptidase A. Maximally tyrosinated and detyrosinated tubulins had similar critical concentrations for polymerization and similar association constants for colchicine binding. Microtubules polymerized from the two tubulins also had similar steady-state mean lengths and length distributions. The growing and shortening dynamics (dynamic instability parameters) of individual microtubules made from maximally tyrosinated or detyrosinated alpha-tubulin as determined by video-enhanced dark-field microscopy were similar, but subtle differences in the growing and shortening rates were found. On balance, however, the dynamicity and thus the overall kinetic stability of the two microtubule populations were indistinguishable. The results support the idea that detyrosination of alpha-tubulin does not by itself generate stable microtubules.
Skoufias
D A
DA
Department of Molecular, Cellular, and Developmental Biology, University of California, Santa Barbara 93106, USA.
Wilson
L
L
D000818:Animals; D002268:Carboxypeptidases / Q000378:metabolism; D043422:Carboxypeptidases A; D002417:Cattle; D003078:Colchicine / Q000378:metabolism*; D019281:Dimerization; D007700:Kinetics; D010453:Peptide Synthases / Q000378:metabolism; D013816:Thermodynamics; D014404:Tubulin / Q000737:chemistry / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism*
D014404:Tubulin; D014443:Tyrosine; D002268:Carboxypeptidases; D043422:Carboxypeptidases A; D010453:Peptide Synthases; C023213:tyrosyltubulin ligase; D003078:Colchicine