A single bifunctional enzyme, PilD, catalyzes cleavage and N-methylation of proteins belonging to the type IV pilin family.

Abstract

Precursors of the type IV pilins of a number of bacterial pathogens, as weil as related proteins involved in extraceliular protein export and DNA uptake, are synthesized with short basic leader sequences. Maturation of these proteins involves two consecutive posttranslational modifications. The leader sequence is fhrst proteolytically removed by specialized endopeptidases, of which the prototype is encoded by the piD gene of Pseudomonas aeruginosa. Subsequently, the amino termini of these proteins are methylated. Here we demonstrate that PilD, in addition to cleaving the amino-terminal leader sequences of prepilin, also catalyzes N-methylation of the amino-terminal phenylalanine of the mature pilin, using S-adenosyl-L-methionine as a methyl donor. Thus, to our knowledge, PilD is the first characterized bacterial N-methyltransferase. Complete inhibition of N-methylation, but not peptide cleavage, by structural analogues of S-adenosyl-L-methionine suggests that PilD is a bifunctional enzyme with proteolytic and methylation activities carried out within two distinct active sites.

Aggregation of heat-shock-denatured, endogenous proteins and distribution of the IbpA/B and Fda marker-proteins in Escherichia coli WT and grpE280 cells

“…It was noticed later, however, that the IbpA/B-immunogen preparation was contaminated by 16 kDa membrane protein. We found that separation of this protein from IbpA/B by one-dimensional electrophoresis could be achieved using a Tricine-containing buffer (Strom et al, 1993). The newly prepared serum does not recognize the 16 kDa membrane protein and does not react with components of the OM fraction.…”

Cell surface localization and processing of the ComG proteins, required for DNA binding during transformation of Bacillus subtilis

“…In addition to the comG proteins, a fourth gene family, exemplified by comC (Mohan et al, 1989), pilD (Strom et al, 1993) and pulO (Dupuy et al, 1992), encodes the peptidases required for processing of pre-pilin-like proteins. Indeed, we have shown that ComGC is processed by a pathway that requires the product of comC (Chung and Dubnau, 1994).…”

BfpU, a soluble protein essential for type IV pilus biogenesis in enteropathogenic Escherichia coli

“…Low-copy-number cloning vector Wang & Kushner (1991) quence and N-methylates the new amino terminus of the pilin (Strom et al, 1993). Pre-pilin peptidases compose a unique class of bi-lobed aspartate proteases (LaPointe & Taylor, 2000).…”