The subunit structure of calf thymus ribonuclease H i as revealed by immunological analysis

Abstract

We have recently reported on the purification, subunit structure, and serological analysis of calf thymus ribonuclease H I and suggested a trimeric or tetrameric structure for the enzyme (Büsen, W., and Vogt, G. (1980) J. Biol. Chem. 255, 9434-9443). Continuation of our immunological analysis, using a protein blotting procedure for antigen detection and immunoaffinity chromatography, revealed that the native enzyme molecule is composed of polypeptides A and C with molecular weights of 31,600 and 24,800 respectively, in a molar ratio of 2 to 1. This is in accordance with a trimeric structure (A,A,C) for calf thymus ribonuclease H I. Polypeptides B and D, found in the most purified fraction, are shown to be generated during the early steps of the purification procedure, suggesting specific protein nicking which does not affect the native molecular weight of the enzyme.