The efficiency of removal of soluble proteins in cell-free bacterial extracts by means of antiserum from rabbits immunized with similar extracts vas measured. Precipitation followed by Sephadex gel-chromatography was ...
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The efficiency of removal of soluble proteins in cell-free bacterial extracts by means of antiserum from rabbits immunized with similar extracts vas measured. Precipitation followed by Sephadex gel-chromatography was used. Up to 807percnt; (exceptionally 907percnt;) removal could be obtained. The method might be applied to enrichment for “foreign” cell-extract components, for example, viral products in virus infected cells. Tests for the specificity of the method are also presented.
An in vitro transcription system for the tryptophan operon of Escherichia coli was used to determine the mechanism by which trp † repressor inhibits transcription of the operon. It was found that trp repressor-binding...
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An in vitro transcription system for the tryptophan operon of Escherichia coli was used to determine the mechanism by which trp † repressor inhibits transcription of the operon. It was found that trp repressor-binding prevents binding of RNA polymerase to the operon. It was also observed that when polymerase is prebound to the trp promoter, repressor cannot prevent it from transcribing. These results suggest the existence of a functional overlap between trp promoter and trp operator regions and indicate that repressor acts by preventing RNA polymerase attachment to the promoter region of the operon.
The lac repressor protein was purified from an Escherichia coli strain carrying an amber mutation in the lacI gene and the tyrosine-inserting amber suppressor, Su3. Protein sequencing showed a change at position 62 in...
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The lac repressor protein was purified from an Escherichia coli strain carrying an amber mutation in the lacI gene and the tyrosine-inserting amber suppressor, Su3. Protein sequencing showed a change at position 62 in the repressor polypeptide chain from leucine to tyrosine, proving that the amber was derived from a UUG codon at this point in the message. This establishes UUG as an initiation codon in vivo , since it has been previously shown that translational reinitiation can occur at position 62.
A soluble protein factor was isolated, free of elongation factor (EF)-T and EF-G, based on its ability to stimulate the synthesis of peptide bonds using ribosomal bound 70S-AUG-N-formyl-[35S]methionyl-tRNA complex and...
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A soluble protein factor was isolated, free of elongation factor (EF)-T and EF-G, based on its ability to stimulate the synthesis of peptide bonds using ribosomal bound 70S-AUG-N-formyl-[35S]methionyl-tRNA complex and added puromycin as substrates. Over 90% of this activity was found in the ribosome-free cytoplasm of Escherichia coli extracts. Otherfeatures such as molecular weight, purification properties, and catalytic activities distinguish this factor from ribosomal proteins and known activators of translation. The factor requires all components needed for peptide bond synthesis and is inhibited by antibiotics known to specifically block the peptidyl transferase activity of ribosomes. The factor increases the binding affinity of the ribosome for the aminoacyl-tRNA analog puromycin about 10-fold. We suggest that this extraribosomal factor modulates the intrinsic activity of ribosomes to catalyze peptide-bond synthesis, and regard it as a new factor required for peptide chain elongation, which we call EF-P.
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