The Process of Infection with Bacteriophage φX174: XIX. Isolation and Characterization of a Chloramphenicol-resistant Protein from φX-infected Cells

Abstract

A chloramphenicol-resistant, viral-directed protein, associated with the synthesis of progeny RF‡ in cells infected with bacteriophage φX174 has been isolated. Uninfected and φX174-infected Escherichia coli C were labeled, in the presence of 30 μg/ml, of CM, with [^(14)C]leucine and [^3H]leucine respectively. The labeled cells were combined, lysed and the lysates fractionated by CaHPO_4 chromatography. A radioactive component (or components) preferentially present in the infected cell extracts was found. This protein was not synthesized in the presence of 100 μg/ml. of CM nor was it formed in nonpermissive cells infected with φX amber mutants of cistron VI; progeny RF is not synthesized under either of these conditions. The CM-resistant φX protein was further separated from contaminant E. coli CM-resistant proteins (14C-labeled) by gel fiitration on Sephadex G100. The elution pattern observed with Sephadex G100 was dependent on the pH and ionic strength. Lysis of the cells with lysozyme, EDTA and Brij, a non-ionic detergent, followed by chromatography on CaHPO_4, yielded more homogeneous preparations than lysis with lysozyme, EDTA and freeze-thawing.

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