Evolution of gram-negative bacteria: a view from lipid A biosynthesis enzymes

Lipid-A, a complex glycolipid, is the highly immunoreactive endotoxic center of lipopolysaccharide (LPS). It anchors the LPS into the outer membrane of most gram-negative bacteria. Lipid A can be recognized by animal cells, triggers some defense-related responses, and causes gram-negative sepsis. The lipid-A biosynthesis pathway consists of nine enzymatic steps. The enzymes involved are: LpxA, LpxC, LpxD, LpxH, LpxB, LpxK, WaaA, LpxL, and LpxM. Traditionally bacteria are classified into two major groups as gram-positive and gram-negative based on Gram-staining procedure. Gram-negative bacteria have LPS-containing outer membranes; hence all these bacteria are expected to possess all genes encoding lipid-A biosynthesis-pathway enzymes. These genes, on the other hand, are likely to be missing from gram-positive bacteria. In order to elucidate how the lipid-A biosynthesis pathway has evolved in the bacterial kingdom, we have examined the distribution of the lipid-A biosynthesis enzymes across 62 bacteria genomes (from 16 phyla including five species of gram-positive bacteria as control). Similarity search was performed using BLASTP and TBLASTN, and also profile hidden Markov models. We started our search using the nine enzyme sequences obtained from the Escherichia coli K12 genome as queries against the other 61 bacterial genomes. We repeated reciprocal searches against all genomes using each of newly identified protein sequences.