Roughly 25-30% of the genes in sequenced genomes are predicted to encode integral membrane proteins. The functions of many of these genes, even in a well-studied organism such as Escherichia coli (E. coli), remain unknown. Two highly conserved and related E. coli inner membrane proteins, YqjA and YghB, are a major interest of the Doerrler lab. They belong to a large membrane protein family (commonly called the DedA family) found widespread in most sequenced bacterial genomes. YghB and yqjA encode inner membrane proteins with 61% amino acid identity. In addition, E. coli contains six additional genes predicted to encode proteins with similarity to YqjA and YghB (yabI, yohD, dedA, ydjX, ydjZ, yqaA). Currently, there are >2000 genes in the NCBI protein database annotated as either belonging to this family or possessing significant amino acid identity to the DedA family (protein BLAST E values < 0.02).
Individually, yghB and yqjA are nonessential genes, as each single deletion mutant grows normally. However, BC202, an E. coli strain with targeted nonpolar deletions of both yqjA and yghB, does not grow at 42 °C and displays a dramatic cell division phenotype by forming chains of cells when grown at the permissive temperature of 30 °C. Phase contrast and scanning electron microscopy analysis of BC202 suggests that mutants can begin cell division but are blocked at a later step in the process.
We have found that the cell division defect of BC202 is due to inefficient export of key periplasmic enzymes called amidases, namely AmiA and AmiC. These enzymes that cut peptidoglycan and are required for the completion of cell division, are normally exported from the cell to the periplasm using a specialized protein export pathway call the twin arginine transport, or Tat, pathway. We demonstrated this pathway functions inefficiently in BC202. The molecular details of this observation remains to be elucidated.
Wild type Escherichia coli (above) BC202 (DyqjA::tet; DyghB::kan)
The dedA homologue of the Lyme Disease pathogen, Borrelia burgdorferi is an essential gene and depletion of the gene product results in cell death preceded by membrane and cell division defects (see photos below, noting the elongated cells that form when the cells are depleted of the plasmid encoded dedA gene; bb0250). These effects are independent of any role this protein family may play in the TAT pathway because the genome of B. burgdorferi does not encode a TAT pathway or any genes with predicted TAT dependent signal peptides.
Borrelia burgdorferi BB0250 mutant (+IPTG) Borrelia burgdorferi BB0250 mutant (-IPTG)