Purification of Xenopus laevis mitochondrial RNA polymerase and identification of a dissociable factor required for specific transcription.
The Xenopus laevis mitochondrial RNA (mtRNA) polymerase was purified to near homogeneity with an overall yield approaching 50%. The major polypeptides in the final fraction were a doublet of proteins of approximately 140 kilodaltons that copurified with the mtRNA polymerase activity. It appeared likely that the smaller polypeptide is a breakdown product of the larger one. The highly purified polymerase was active in nonspecific transcription but required a dissociable factor for specific transcription of X. laevis mtDNA. The factor could be resolved from mtRNA polymerase by hydrophobic chromatography and had a sedimentation coefficient of 3.0 S. The transcription factor eluted from both the hydrophobic column and a Mono Q anion-exchange column as a single symmetrical peak. The mtRNA polymerase and this factor together are necessary and sufficient for active transcription from four promoters located in a noncoding region of the mtDNA genome between the gene for tRNA(Phe) and the displacement loop.
Bogenhagen
D F
DF
Department of Pharmacology, State University of New York, Stony Brook 11794.
Insdorf
N F
NF
D000818:Animals; D002845:Chromatography; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D004272:DNA, Mitochondrial / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000302:isolation & purification*; D012313:RNA / Q000378:metabolism; D000077278:RNA, Mitochondrial; D014157:Transcription Factors / Q000302:isolation & purification*; D014158:Transcription, Genetic; D014982:Xenopus laevis
D004272:DNA, Mitochondrial; D000077278:RNA, Mitochondrial; D014157:Transcription Factors; D012313:RNA; D012321:DNA-Directed RNA Polymerases