Purification and characterization of tubulin-tyrosine ligase from porcine brain.
A tubulin-tyrosine ligase was purified from porcine brains using DEAE-cellulose chromatography, Sepharose-sebacic acid hydrazide-ATP affinity chromatography and Sepharose-tubulin affinity chromatography. The purified enzyme migrated as a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and its molecular weight was estimated to be 46,000. The apparent molecular weight was 37,000 on a high speed liquid chromatograph equipped with gel filtration columns. The pH optimum for the activity was around 8 and a second peak was observed at around 6.5 8.5 microM ATP or 30 microM tyrosine gave half-maximal activity. The purified enzyme catalyzed the tyrosination of the alpha subunit of tubulin in vitro.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001921:Brain / Q000201:enzymology*; D055598:Chemical Phenomena; D002621:Chemistry; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D010453:Peptide Synthases / Q000302:isolation & purification*; D013552:Swine; D014404:Tubulin / Q000302:isolation & purification; D014443:Tyrosine / Q000302:isolation & purification
D000596:Amino Acids; D014404:Tubulin; D014443:Tyrosine; D010453:Peptide Synthases; C023213:tyrosyltubulin ligase