Type IV prepilin peptidase gene of Neisseria gonorrhoeae MS11: presence of a related gene in other piliated and nonpiliated Neisseria strains

Abstract

The assembly of type IV pili in Neisseria gonorrhoeae is a complex process likely to require the products of many genes. One of these is the enzyme prepilin peptidase, which cleaves and then N methylates the precursor pilin subunits prior to their assembly into pili. We have used a PCR amplification strategy to clone the N. Pili are important virulence factors for certain types of human bacterial pathogens, such as Pseudomonas aeruginosa (9, 67), Vibrio cholerae (64), and Neisseria gonorrhoeae (43, 61). These three species all produce type IV pili composed of a single major protein subunit (pilin). These are produced as precursors which are processed at a highly conserved consensus cleavage site [G l F(M)TLXE] located close to the amino terminus (12,55, 58). Characteristically, the mature pilins have an N-methylated amino-terminal residue (usually phenylalanine or methionine) and retain a long continuous segment of hydrophobic amino acids close to their N terminus which is essential for their polymerization (19,53, 58).The assembly of type IV pili has been most extensively studied in P. aeruginosa, in which four assembly proteins (PilB, PilC, and PilD [37] and PilQ [29]) have been identified, and in V cholerae, in which at least seven assembly proteins (TcpB,41,44]) have been identified. Although the roles of most of these proteins remain obscure, PilD from P. aeruginosa (PilDpa) and TcpJ are known to correspond to the prepilin peptidase, which cleaves the pilin precursors (22,38), and PilD has additionally been shown to N methylate the processed product (60). Intriguingly, several bacteria which are not known to have type IV pili also produce a protein with significant sequence homology to known prepilin peptidases and which is required for extracellular secretion (45). One of these proteins, PulO of Klebsiella oxytoca, can correctly cleave and N methylate the precursor of gonococcal type IV pilin (11) as well as the precursor of a type IV pilin-like protein (PulG) of K oxytoca (46,47 Relatively little is known about the assembly of type IV pili in N. gonorrhoeae. Expression of the N. gonorrhoeae, Dichelobacter nodosus, or Moraxella bovis type IV pilin genes in P. aeruginosa results in their assembly into pili (13,20,31), suggesting that the mechanism of type IV pilus assembly is basically the same in all of these bacteria. One of the major difficulties encountered in attempts to use genetic techniques to identify factors necessary for pilus formation in N. gonorrhoeae is the high frequency of pilus phase variation caused by genome rearrangements (33). However, Jonsson et al. (21) have identified a gene (N. gonorrhoeae pilC [PlCNg]) which seems to be required for piliation, although this view has recently been challenged by Rudel et al. (51), who demonstrated that PilCNg might be required for pilus-mediated adherence rather than for pilus assembly.One of the aspects of N. gonorrhoeae piliation which seems to be readily amenable to genetic analysis is the processing of the pilin precursor. All type IV pilin...

General secretion pathway (eps) genes required for toxin secretion and outer membrane biogenesis in Vibrio cholerae

“…Analysis of the DNA se- quence 600 bp downstream of epsN did not reveal any ORFs with homology to known GSP genes but did identify a partial ORF, in the opposite orientation from epsN, with similarity to the cysQ gene of E. coli (46). This is in contrast to most other GSP operons, in which an additional gene, encoding a prepilin peptidase required for secretion, is present directly downstream of the epsN gene (3,12,55).…”

“…This is different from most other GSP operons, in which a prepilin peptidase gene is present directly downstream of the -N gene (3,12,55). However, it demonstrates the similarity of the V. cholerae GSP system to GSP systems in P. aeruginosa and Aeromonas hydrophila, in which the peptidase gene is found elsewhere on the chromosome.…”

A plasmid-encoded prepilin peptidase gene from enteropathogenic Escherichia coli

“…Previous reports have described the ability of prepilin peptidases from K oxytoca, B. subtilis, and P. aeruginosa to process the prepilin of N. gonorrhoeae (11,35). In addition, the gonococcal prepilin peptidase has been shown to process the PulG protein of K oxytoca (10), and the enzyme from P. aeruginosa has been shown to process the ExeG protein of A. hydrophila (20). However, the TcpJ protein of V cholerae is not able to process the prepilin of P. aeruginosa when both are cloned in E. coli (22).…”

PpdD Type IV Pilin of Escherichia coli K-12 Can Be Assembled into Pili in Pseudomonas aeruginosa

“…The precise role of each piliation protein and the way their genes are regulated are unclear. However, it is known that type IV pili contain a major pilin subunit that is proteolytically processed and N methylated by a prepilin peptidase (13,18,33,35). This enzyme, together with the major and minor pilin subunits (2), an outer membrane protein (secretin) (6,11,22) with its chaperone-like protein (12), and an ATP binding protein (37) are the best-characterized components of the type IV piliation machinery.…”