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A mutation in thelon (capR ) gene ofEscherichia coli K-12 results in overproduction of capsular polysaccharide and increased sensitivity to ultraviolet and ionizing radiations. Thelon (capR ) gene deoxyribonucleic acid was cloned from a new F′ factor. The new plasmids, designated pBZ201 and pBZ203, (i) contained an additional 8.2-megadalton (Md)Eco RI fragment that had the same mobility as one of theEco RI fragments of the F′, and (ii) conferred repression of capsular polysaccharide synthesis and repression of sensitivity to ultraviolet radiation in a bacterial transformation experiment withcapR mutant recipient strains. AcapR9 mutant plasmid, pBZ201M9, was also isolated and conferred expression of mucoidy and ultraviolet sensitivity to acapR + (lon + ) strain, indicating that thecapR9 allele was dominant. Plasmids pBZ201M80, pBZ201M9-INSA, and pBZ201M9-INSB were characterized by transformation as containing recessivecapR mutant alleles. Heteroduplex analyses and agarose gel electrophoresis of restriction endonuclease digests of plasmid DNA preparations revealed that (i) pBZ201M9-INSA and pBZ201M9-INSB each contains a 0.5-Md insertion (probably IS1 ) in the cloned DNA fragment at the same site, and (ii) pBZ201 and pBZ203, bothcapR + plasmids, contain the same 8.2-Md fragment cloned in opposite orientations with respect to the cloning vehicle, pSC101. Plasmid-specified polypeptides were determined by using strain CSR603 maxicells containing each plasmid. Two new polypeptides were coded by thelon + (capR + ) 8.2-Md DNA fragment: Z1, 94 kilodaltons (94K), and Z2, 67K. The maxicells containing recessivecapR mutant plasmids were deficient only in synthesis of the 94K polypeptide, and the dominant (capR9 ) mutant plasmid specified 5 to 10 times more of the 94K polypeptide than the maxicells containing thecapR + plasmid. Other data indicated that thecapR9 -specified “94K polypeptide” was not identical to thecapR + -specified “94K polypeptide.” Thus the altered mutant polypeptide was synthesized in increased quantities, suggesting a defective mode of autogenous regulation for thecapR9 polypeptide and effective autogenous regulation of thecapR + polypeptide.

journal of bacteriology

Cloning of Gene lon ( capR ) of Escherichia coli K-12 and Identification of Polypeptides Specified by the Cloned Deoxyribonucleic Acid Fragment