Identification of the dye gene product, mutational loss of which alters envelope protein composition and also affects sex factor F expression in Escherichia coli K-12

Summary

The product of the dye gene of Escherichia coli, mapping at 99–100 min, is required for expression of the sex factor F, and also appears to be involved in the regulation of envelope proteins. Mutation of dye thus results in loss of expression of the F-factor (Fex⁻, i.e. male sterility, and dye sensitivity (Dye^s). We have isolated a plasmid, pRB38, in which a 6 kb SalI fragment carrying the dye ⁺ gene was cloned into the plasmid pACYC184. This 6 kb SalI fragment also carries two nearby markers, chlG, involved in the synthesis of the molybdenum cofactor, and phoM, required for constitutive expression of alkaline phosphatase.

Some of the polypeptides synthesised by pRB38 were identified using the maxi-cell procedure. The product of the dye gene was found to be a polypeptide of M_r=29,000. Thus derivatives of pRB38 in which the transposon γδ was inserted into dye, resulting in a Dye^S Fex⁻ phenotype when these plasmids were in a Δ dye strain, failed to produce this polypeptide and in some cases produced a truncated product. Such insertions also resulted in a Chl^r and Pho⁻ phenotype when the plasmid was in a Δ(dye-chlG-phoM) phoR strain, although complementation tests suggested that the phoM ⁺ and chlG ⁺ genes were still intact. Insertions of γδ into the promoter distal end of dye did not result in a Dye^S Fex⁻ phenotype, although a truncated Dye protein was synthesised, and a Chl^r Pho⁻ phenotype was produced.

It has been suggested (Gaffney et al. 1983) that the dye (=sfrA) gene product is necessary for F-factor expression because it is required for translocation of the F-factor TraJ protein to the outer membrane. Our results suggest that the Dye protein is also required for expression of the molybdenum cofactor and of alkaline phosphatase, and could perhaps be involved in the translocation of these proteins to the membrane.