Purification and subunit structure studies of human placental threonyl-tRNA synthetase.

Human threonyl-tRNA synthetase has been purified from full-term placenta over 5000-fold with a 13% overall yield by a combination of affinity chromatographic methods and conventional procedures. The product was apparently homogeneous by the criterion of sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The purified enzyme catalyzed the esterification of approximately 3500 nmol L-threonine to tRNA per min per mg of protein at 37 degrees, corresponding to a molecular activity of approximately 700/min. The apparent molecular weight of the native enzyme ranged from 210000 to 220000 by gel-filtration on Sephadex G-200, and was either approximately 110000 or 228000 by sucrose gradient centrifugation for different preparations. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis demonstrated a single band of molecular weight 85000 or 115000 for different preparations. These results suggest that human placental threonyl-tRNA synthetase has a subunit structure of the type alpha 2 with a subunit molecular weight of about 100000. However, some other molecular forms with lower specific activity were also found to exist under certain conditions.