Purification and properties of the polypeptide chain release factor (RF-4) from an extreme thermophile, Thermus thermophilus HB8.
The polypeptide chain release factor 1 (RF-1) has been purified from an extreme thermophile, Thermus thermophilus HB8. The purification procedure included steps of aqueous two-phase partition, ammonium sulfate fractionation, and column chromatographies on DEAE-Sephadex, Sephadex G-150, and CM-Sephadex. The preparation was more than 90% pure as judged by polyacrylamide gel electrophoresis. The specific activity was about 3.3 pmol of formyl-[3H]-methionine released in 1 min at 25 degrees C per microgram of protein under the standard assay conditions using 4 pmol of the initiation complex and 1 nmol of UpApG. The molecular weight as determined by gel filtration on Sephadex G-150 and by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, was 58,000 and 45,000, respectively. As expected, the factor was extremely heat-stable, 50% of its activity remaining after incubation for 5 min at 84 degrees C. Several properties of the reaction catalyzed by RF-1 are also described.
D001426:Bacterial Proteins / Q000302:isolation & purification / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D005033:Ethylmaleimide / Q000494:pharmacology; D007700:Kinetics; D009239:N-Formylmethionine / Q000378:metabolism; D009843:Oligoribonucleotides / Q000494:pharmacology; D010443:Peptide Chain Termination, Translational / Q000187:drug effects; D010454:Peptide Termination Factors / Q000302:isolation & purification; D012270:Ribosomes / Q000378:metabolism; D013824:Thermus / Q000378:metabolism*
D001426:Bacterial Proteins; D009843:Oligoribonucleotides; D010454:Peptide Termination Factors; D009239:N-Formylmethionine; D005033:Ethylmaleimide