DEAD DNA Transport: a mutational analysis of a competence helicase/translocase

Abstract

In bacteria, horizontal gene transfer (HGT) is a major mechanism for generating genetic diversity. The predominant HGT processes all involve the transfer of large macromolecules across cell membranes. Genetic competence is one such process in which cells are able to take up DNA from their environment in a processive manner. These cells become transformed when the acquired genes are maintained a part of the organism's genome or as an extra‐chromosomal element such as a plasmid. Many bacteria carry homologs of the central DNA uptake machinery that has been well characterized in Bacillus subtilis. The core machinery is comprised of a DNA binding receptor (ComEA), an aqueous pore (ComEC), and a helicase/translocase (ComFA). It has been postulated that ComFA belongs to a family of low processivity helicases/translocases, the DEAD‐box family. Thus, it is currently unclear how ComFA contributes to the high processivity exhibited during DNA uptake. In the work presented here, we conduct a mutational analysis of ComFA based upon sequence homology to conserved DEAD‐box helicase/translocase motifs. Mutating the conserved residues disrupts DNA uptake and transformation, suggesting that ComFA is in fact a member of the DEAD‐box family. Understanding how the uptake machinery functions expands our understanding of the diversity of cellular functions that employ DEAD‐box helicases, and possibly the diversity of behaviors of the protein family.