Residual polyadenylation in poly(A) polymerase I (pcnB) mutants of Escherichia coli does not result from the activity encoded by the f310 gene

As extracts of poly(A) polymerase I(PAP I) deficient strains of Escherichia coil appeared to contain considerable residual polyadenylating activity, e fforts were undertaken to identify a second poly(A) polymerase. Recently, a gene (f310) encoding the putative second poly(A) polymerase was cloned and sequenced. Here we have tested the ability of the F310 protein to add poly (A) tails in vivO by measuring total poly(A) levels in both f310 mutants an d strains that overproduce F310. In addition, we have visualized poly(A) ta ils and examined ColE1 plasmid copy number in various genetic backgrounds. We also carried out direct biochemical measurements of AMP incorporation, u sing cell extracts after amplification of F310. All the data obtained indic ate that F310 is not a poly(A) polymerase. Although the presence of two pot ential ATP binding domains in the F310 protein may account for its apparent ATP binding activity, its true biochemical function remains to be identifi ed. In addition, we show that the f310 gene is transcribed, almost exclusiv ely, during stationary phase from a sigma(S) promoter.