Xenopus transcription factor IIIA-dependent DNA renaturation.
Kinetic and titration analyses are used to elucidate the mechanism by which Xenopus transcription factor IIIA (TFIIIA), a protein required for 5 S RNA synthesis by RNA polymerase III, promotes DNA renaturation. TFIIIA promotes 50% renaturation of complementary strands (303 bases) in 45 s. Analyses of the renaturation kinetics indicate the rate-limiting step in this TFIIIA-dependent reaction is first order. TFIIIA-dependent DNA renaturation is a stoichiometric rather than a catalytic process. The renaturation rates for specific and nonspecific DNA are very similar, indicating lack of sequence specificity in this TFIIIA-dependent process. In the nanomolar concentration range of protein and DNA, renaturation occurs at a ratio of about one TFIIIA molecule/single strand (303 bases). Elevated reaction temperatures strongly stimulate TFIIIA-dependent DNA renaturation; at 45 degrees C, renaturation of the 303-base pair fragment nears completion in about 5 s. The ability of TFIIIA to rapidly promote DNA renaturation is unique when compared with Escherichia coli recA protein, single-stranded DNA binding protein, or bacteriophage T4 gene 32 protein. This mechanism by which TFIIIA promotes DNA renaturation is compatible with features of 5 S RNA gene transcription.
Fiser-Littell
R M
RM
Department of Biochemistry and Molecular Biology, University of Oklahoma Health Sciences Center, Oklahoma City 73190.
Hanas
J S
JS
D000818:Animals; D004247:DNA / Q000032:analysis*; D004268:DNA-Binding Proteins / Q000378:metabolism; D007700:Kinetics; D009695:Nucleic Acid Renaturation; D012313:RNA / Q000096:biosynthesis; D013696:Temperature; D035582:Transcription Factor TFIIIA; D014157:Transcription Factors / Q000378:metabolism*; D014982:Xenopus laevis
D004268:DNA-Binding Proteins; D035582:Transcription Factor TFIIIA; D014157:Transcription Factors; D012313:RNA; D004247:DNA