Purification and subunit structure studies of human placental threonyl‐tRNA synthetase

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Abstract
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 ˜ 3500 nmol L‐threonine to tRNA per min per mg of protein at 37°, corresponding to a molecular activity of ˜ 700/min. The apparent molecular weight of the native enzyme ranged from 210 000 to 220 000 by gel‐filtration on Sephadex G‐200, and was either ˜ 110 000 or 228 000 by sucrose gradient centrifugation for different preparations. Sodium dodecyl sulfate‐polyacrylamide gel electrophoresis demonstrated a single band of molecular weight 85 000 or 115 000 for different preparations. These results suggest that human placental threonyl‐tRNA synthetase has a subunit structure of the type α2 with a subunit molecular weight of about 100 000. However, some other molecular forms with lower specific activity were also found to exist under certain conditions.