Polypeptides of DNA-dependent RNA polymerase of spinach chloroplasts: characterization by antibody-linked polymerase assay and determination of sites of synthesis.

Using solid-phase ;Sandwich' immunoassays we studied DNA-dependent RNA polymerase of spinach chloroplasts with regard to (i) polypeptide composition of the multimeric enzyme; (ii) immunological cross-reaction with Escherichia coli RNA polymerase; (iii) sites of synthesis of polymerase polypeptides. Our main results are as follows. (i) All polypeptides of isolated chloroplast RNA polymerase (150, 145, 110, 102, 80, 75 and 38 kd) are labeled by an antibody-linked polymerase assay (ALPA), i.e., they are immunologically related to subunits of the holoenzyme. On the other hand differences in the patterns of ;ALPA-reactive' polypeptides of a crude RNA polymerase fraction and of the purified enzyme preparation indicate partial proteolytic degradation of polymerase polypeptides during purification. Thus the 80- and 75-kd polypeptides, which had been previously considered as true RNA polymerase polypeptides, probably result from partial proteolytic degradation. (ii) The 150- and 145-kd polypeptides show immunochemical similarities with the beta and/orbeta' subunits of E. coli RNA polymerase. (iii) Results from solidphase immunoassay of in vitro translated products of both chloroplast RNA and poly(A) (nuclear) RNA suggest that all chloroplast RNA polymerase polypeptides are coded for by the nucleus.