Characterization of the post-translational modifications in tubulin from the marginal band of avian erythrocytes.
Tubulin purified from turkey erythrocytes was characterized by partial protein sequence data, high-resolution IEF and by its reaction with antibodies specific for certain post-translational modifications. The tubulin from the marginal band contains a single alpha and beta isotype, i.e. alpha 1 and beta 6. Partial protein sequences and immunoblotting with antibody 6-11B-1 show that erythrocyte alpha 1 tubulin is not acetylated at Lys40. The acidic carboxy-terminal peptides purified by Mono Q chromatography and reverse-phase HPLC were characterized by sequence analysis and mass spectrometry. Although erythrocyte alpha tubulin is almost completely detyrosinated it retains the penultimate glutamic acid residue, which is partially lost in brain tubulin. Thus erythrocyte tubulin is an excellent substrate for extensive in vitro tyrosination by tubulin-tyrosine ligase. Erythrocyte alpha and beta tubulin lack the side-chain polyglutamylation found in all major tubulins from adult brain. Finally we show that about 10% of the beta tubulin is phosphorylated at Ser441. Thus erythrocyte tubulin is an unusual homogeneous preparation. It contains the minimum possible number of tubulin isotypes and the only post-translational modifications detected (detyrosination and phosphorylation) are reversible.
Rüdiger
M
M
Max Planck Institute for Biophysical Chemistry, Department of Biochemistry, Göttingen, Germany.
Weber
K
K
D000595:Amino Acid Sequence; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D004912:Erythrocytes / Q000378:metabolism*; D007525:Isoelectric Focusing; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010453:Peptide Synthases / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D013552:Swine; D014404:Tubulin / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D014422:Turkeys; D014443:Tyrosine / Q000378:metabolism
D014404:Tubulin; D014443:Tyrosine; D010453:Peptide Synthases; C023213:tyrosyltubulin ligase