Open AccessStructural Relationships Among the Multiple Forms of DNA‐Dependent RNA Polymerase II from Cultured Parsley Cells
Author(s) -
LINK Gerhard,
KIDD George H.,
BOGORAD Lawrence
Publication year - 1978
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1978.tb12688.x
Subject(s) - polymerase , microbiology and biotechnology , rna polymerase , protein subunit , trypsin , rna , gel electrophoresis , biology , enzyme , biochemistry , polyacrylamide gel electrophoresis , chemistry , gene
DNA‐dependent RNA polymerase II (or B) was purified from cultured parsley cells, and its molecular structure was examined in detail. Upon centrifugation through glycerol gradients, RNA polymerase II sediments as a single band with an apparent sedimentation constant of 15 S. No contamination with RNA polymerases I or III could be detected when the activity of purified RNA polymerase II was assayed in the presence of high concentrations of α‐amanitin. Analysis of purified RNA polymerase II by nondenaturing and denaturing polyacrylamide gel electrophoresis revealed that this enzyme exists in multiple forms. They were designated II o , II A , and II B . It is suggested that each form has a subunit of M r = 140 000 as well as smaller polypeptides in common. They differ, however, in the molecular weights of their largest subunits which is 220 000 in form II o , 200 000 in form II A , and 180 000 in form II B . These large subunits were labelled with 125 I, digested with trypsin, and tryptic digests were compared by two‐dimensional analysis on thin‐layer plates [Elder et al. (1977) J. Biol. Chem. 252 , 6510–6515]. Fingerprints of tryptic digests from the polypeptides with M r = 220 000, M r = 200 000, and M r = 180 000 were similar. It is, therefore, suggested that these subunits are structurally related. A tryptic digest was also produced from the subunit with M r = 140 000. Its fingerprint was found to yield a considerably different distribution of peptides as compared to those from the three large subunits.