Recognition of specific Physarum alpha-tubulin isotypes by a monoclonal antibody. Sequence heterogeneity around the acetylation site at lysine 40.
The monoclonal antibody 6-11B-1 recognises specifically the acetylated form of alpha-tubulin. The acetylation event occurs on a unique lysine residue, lysine 40. Using 6-11B-1, acetylated alpha-tubulin was detected in myxamoebae but not plasmodia of Physarum polycephalum. Following chemical acetylation plasmodial alpha-tubulin was detected by 6-11B-1. The monoclonal antibody KMP-1 recognises certain Physarum alpha-tubulin isotypes but only in non-acetylated form. Whilst recognising all the non-acetylated fraction of myxamoebal alpha-tubulin only a proportion of plasmodial alpha-tubulin was recognised by KMP-1. Peptides were synthesised corresponding to the acetylation domains (containing lysine 40) of myxamoebal alpha-tubulin and the inferred acetylation domains of two plasmodial-specific alpha-tubulin isotypes. The only difference between the two peptides was at a single residue corresponding to amino acid 44 in the polypeptide. Tyrosine was present in myxamoebal alpha-tubulin and glycine was present in the plasmodial specific peptides; the peptides are referred to as the Tyr44 and Gly44 peptides respectively. Both peptides in acetylated form blocked 6-11B-1 reactivity towards acetylated myxamoebal alpha-tubulin. The Tyr44 but not the Gly44 peptide blocked KMP-1 reactivity towards non-acetylated myxamoebal alpha-tubulin. Tyrosine at position 44 is not found in any other known alpha-tubulin. Thus a unique antigenic determinant exists in certain Physarum alpha-tubulin isotypes, close to the acetylation site at lysine 40. This antigenic determinant forms part of the KMP-1 recognition epitope and explains the unique isotype selectivity of this monoclonal antibody.
Walden
P D
PD
Biological Laboratory, University of Kent, Canterbury, England.
Blindt
A B
AB
Birkett
C R
CR
Cox
R A
RA
Gull
K
K
D000107:Acetylation; D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology; D000918:Antibody Specificity; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000276:immunology*; D015151:Immunoblotting; D007527:Isoenzymes / Q000276:immunology; D008239:Lysine / Q000378:metabolism; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D010455:Peptides / Q000138:chemical synthesis / Q000378:metabolism; D010804:Physarum / Q000378:metabolism*; D014404:Tubulin / Q000276:immunology* / Q000378:metabolism
D000911:Antibodies, Monoclonal; D000939:Epitopes; D007527:Isoenzymes; D010455:Peptides; D014404:Tubulin; D008239:Lysine