Glycosylation of yeast aspartyl-tRNA synthetase. Affinity labelling by glucose and glucose 6-phosphate.

Several lines of evidence establish that the crystallizable aspartyl-tRNA synthetase from Baker's yeast contains some covalently bound glucose: (i) a positive staining of the enzyme was obtained after polyacrylamide gel electrophoresis followed by the concanavalin A-peroxidase test which is specific for glucose and mannose containing proteins; (ii) thin-layer chromatography and gas-liquid chromatography revealed the presence of glucose in enzyme hydrolysates; (iii) immunoaffinoelectrophoresis in agarose gels containing concanavalin A and antibodies raised against aspartyl-tRNA synthetase showed that the enzyme was able to precipitate entirely in the lectin. Finally incubation of the enzyme with [14C]glucose or [14C]glucose 6-phosphate led to the incorporation of radioactivity into trichloroacetic acid-precipitable protein. Indeed immunoprecipitation of [14C]glucose-labelled aspartyl-tRNA synthetase with specific antibodies using the rocket method followed by autoradiography gave a radioactive peak. This last result also demonstrates the possibility of in vitro glycosylation of yeast aspartyl-tRNA synthetase.