Identification of an acetylation site of Chlamydomonas alpha-tubulin.
An acetylation site of Chlamydomonas axonemal alpha-tubulins was identified near, or within, the binding site of 6-11B-1, a monoclonal antibody specific for posttranslationally acetylated alpha-tubulins. In a first approach, axonemal proteins were hydrolyzed by formic acid, cyanogen bromide, or chymotrypsin and analyzed with immunoblots. The smallest alpha-tubulin peptide retained on nitrocellulose and containing antibody-binding site(s) was found to span amino acids 37-138 (alpha 37-138). A smaller antibody-binding peptide, identified as alpha 25-50, was obtained by complete digestion of alpha-tubulin with chymotrypsin. This fragment was purified by reversed-phase HPLC and assayed by its ability to bind 6-11B-1 in solution. Determination of the amino acid sequences of alpha 37-138 and alpha 25-50 showed that residue 40 in axonemal alpha-tubulin is epsilon N-acetyllysine. A sequence very similar to Chlamydomonas alpha 25-50 is found in the majority of alpha-tubulins analyzed so far. However, the corresponding region is markedly divergent in some alpha-tubulin isoforms from chicken, Drosophila, and yeast.
D000107:Acetylation; D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D001665:Binding Sites; D002696:Chlamydomonas / Q000032:analysis*; D000939:Epitopes / Q000032:analysis; D010449:Peptide Mapping; D011499:Protein Processing, Post-Translational; D014404:Tubulin / Q000032:analysis*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D014404:Tubulin