Suppression of amber mutants in vitro induced by low temperature. |
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Authors: | J L Manley R F Gesteland |
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Institution: | Cold Spring Harbor Laboratory Cold Spring Harbor N.Y. 11724, U.S.A. |
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Abstract: | Amber mutations are efficiently and specifically suppressed during protein synthesis in vitro in an Su? S-30 extract at 25 °C, but not at 37 °C. Eight different amber mutations in three different genes have been tested, and all are suppressed. The efficiencies of suppression range from 20 to 35%, when protein synthesis is at the Mg2+ concentration optimal for β-galactosidase synthesis at 25 °C. The suppression efficiency increases to approximately 60% at higher Mg2+ concentrations, and is reduced to less than 5% at very low concentrations. Ochre and UGA mutations are not suppressed at all under these conditions. The amber suppression is inhibited by addition of a purified protein synthesis release factor to the reaction, or when the protein synthesis reaction takes place in extracts derived from bacteria which are streptomycin-resistant. |
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