Peptidoglycan LD-Carboxypeptidase Pgp2 Influences Campylobacter jejuni Helical Cell Shape and Pathogenic Properties, and Provides the Substrate for the DL-Carboxypeptidase Pgp1

Despite the importance of Campylobacter jejuni as a pathogen, little is known about the fundamental aspects of its peptidoglycan (PG) structure and factors modulating its helical morphology. A PG dl-carboxypeptidase Pgp1 essential for maintenance of C. jejuni helical shape was recently identified. Bioinformatic analysis revealed the CJJ81176_0915 gene product as co-occurring with Pgp1 in several organisms. Deletion of cjj81176_0915 (renamed pgp2) resulted in straight morphology, representing the second C. jejuni gene affecting cell shape. The PG structure of a Δpgp2 mutant showed an increase in tetrapeptide-containing muropeptides and a complete absence of tripeptides, consistent with ld-carboxypeptidase activity, which was confirmed biochemically. PG analysis of a Δpgp1Δpgp2 double mutant demonstrated that Pgp2 activity is required to generate the tripeptide substrate for Pgp1. Loss of pgp2 affected several pathogenic properties; the deletion strain was defective for motility in semisolid agar, biofilm formation, and fluorescence on calcofluor white. Δpgp2 PG also caused decreased stimulation of the human nucleotide-binding oligomerization domain 1 (Nod1) proinflammatory mediator in comparison with wild type, as expected from the reduction in muropeptide tripeptides (the primary Nod1 agonist) in the mutant; however, these changes did not alter the ability of the Δpgp2 mutant strain to survive within human epithelial cells or to elicit secretion of IL-8 from epithelial cells after infection. The pgp2 mutant also showed significantly reduced fitness in a chick colonization model. Collectively, these analyses enhance our understanding of C. jejuni PG maturation and help to clarify how PG structure and cell shape impact pathogenic attributes.