Pepsin homologues in bacteria

Abstract

Background: Peptidase family A1, to which pepsin belongs, had been assumed to be restricted to eukaryotes. The tertiary structure of pepsin shows two lobes with similar folds and it has been suggested that the gene has arisen from an ancient duplication and fusion event. The only sequence similarity between the lobes is restricted to the motif around the active site aspartate and a hydrophobic-hydrophobic-Gly motif. Together, these contribute to an essential structural feature known as a psi-loop. There is one such psi-loop in each lobe, and so each lobe presents an active Asp. The human immunodeficiency virus peptidase, retropepsin, from peptidase family A2 also has a similar fold but consists of one lobe only and has to dimerize to be active. All known members of family A1 show the bilobed structure, but it is unclear if the ancestor of family A1 was similar to an A2 peptidase, or if the ancestral retropepsin was derived from a half-pepsin gene. The presence of a pepsin homologue in a prokaryote might give insights into the evolution of the pepsin family.

Results: Homologues of the aspartic peptidase pepsin have been found in the completed genomic sequences from seven species of bacteria. The bacterial homologues, unlike those from eukaryotes, do not possess signal peptides, and would therefore be intracellular acting at neutral pH. The bacterial homologues have Thr218 replaced by Asp, a change which in renin has been shown to confer activity at neutral pH. No pepsin homologues could be detected in any archaean genome.

Conclusion: The peptidase family A1 is found in some species of bacteria as well as eukaryotes. The bacterial homologues fall into two groups, one from oceanic bacteria and one from plant symbionts. The bacterial homologues are all predicted to be intracellular proteins, unlike the eukaryotic enzymes. The bacterial homologues are bilobed like pepsin, implying that if no horizontal gene transfer has occurred the duplication and fusion event might be very ancient indeed, preceding the divergence of bacteria and eukaryotes. It is unclear whether all the bacterial homologues are derived from horizontal gene transfer, but those from the plant symbionts probably are. The homologues from oceanic bacteria are most closely related to memapsins (or BACE-1 and BACE-2), but are so divergent that they are close to the root of the phylogenetic tree and to the division of the A1 family into two subfamilies.

Figure 1

Alignment of bacterial pepsin homologues with human pepsin A and memapsins 1 and 2. Residues are numbered according to mature human pepsin A. Inserts relative to pepsin A are indicated by letters. Each active site residue (Asp32, Tyr75 and Asp215) is indicated by an asterisk. The hydrophobic-hydrophobic-Gly motifs in the psi-loops are indicated by ampersands. A disulphide bridge is indicated by a slash over one cysteine followed by a number and a backslash preceded by the same number over the second cysteine. The Chroma software [37] has been used to highlight residues according to amino acid properties and to generate the consensus line below each alignment block showing 80% conservation or more. Inserts found in only one sequence have been removed and are indicated by the number of amino acids excised in parentheses.

Figure 2

Phylogenetic tree derived from members of peptidase family A1. The tree was generated for all peptidase unit sequences of family A1 holotypes, plus those from the sequences of the bacteria listed in Table 1. The tree is unrooted, but the sequence of plasmepsin-5, which is very divergent, was chosen as the outgroup. Homologues from bacteria are highlighted with a blue background. Branches that were present in 90% of the bootstrap trees are shown in red. Key: [see Additional file 1].