Low molecular weight aspartyl-tRNA synthetase from porcine thyroid. Purification, characterization, and heterogeneity.

The total low molecular weight aspartyl-tRNA synthetase activity of porcine thyroid is distributed among four distinct forms, all of which are identical in size, as determined by gel filtration. The predominant form was purified 25,000-fold to near homogeneity. A high concentration of glycerol (25%, v/v) was required throughout the procedure to maintain stability. The native enzyme was of the alpha 2-type with a Mr = 120,000 estimated by gel filtration. Its subunits were Mr = 53,000 as determined using polyacrylamide gel electrophoresis under denaturing conditions. The enzyme had an isoelectric point of pH 5.4 and pH optimum that varied from pH 7.3 to 8.8 depending on the type of buffer present. The variation in pH optimum was related to a salt effect. All salts tested were inhibitory, with the degree of inhibition dependent on the anion present. Inorganic pyrophosphate was a particularly powerful inhibitor; Km values for aspartate and tRNAAsp were significantly reduced in the presence of inorganic pyrophosphatase. Evidence is presented that the allotropism of the low molecular weight forms is not due to phosphorylation, proteolytic degradation, or stable enzyme-substrate complexes.