TgpA, a Protein with a Eukaryotic-Like Transglutaminase Domain, Plays a Critical Role in the Viability of Pseudomonas aeruginosa

Abstract

The Gram-negative bacterium Pseudomonas aeruginosa is an important opportunistic pathogen in compromised individuals, such as patients with cystic fibrosis, severe burns or impaired immunity. In this work we aimed to screen novel essential genes of P. aeruginosa by shotgun antisense identification, a technique that was developed a decade ago for the Gram-positive bacterium Staphylococcus aureus and was under-used in Gram-negative bacteria for a considerable period of time. Following antisense screenings in the PAO1 strain of P. aeruginosa, we focused on a locus, PA2873, which was targeted by an antisense RNA construct that can impair cell growth. The PA2873 gene product was annotated as a hypothetical membrane protein endowed with a periplasmic region harbouring a structural domain belonging to the transglutaminase-like superfamily, a group of archaeal, bacterial and eukaryotic proteins homologous to animal transglutaminases. In this study, we show that the periplasmic portion of the PA2873 protein, which we named TgpA, does possess transglutaminase activity in vitro. This is the first report of transglutaminase activity in P. aeruginosa. In addition, we have provided strong evidences that TgpA plays a critical role in the viability of P. aeruginosa.

The Identification and Functional Characterization of WxL Proteins from Enterococcus faecium Reveal Surface Proteins Involved in Extracellular Matrix Interactions

“…Sections of DufA and DufC both modeled to the three-dimensional model of the human transglutaminase 32 enzyme with the highest confidence level (98.8%), using 59% and 52% of the protein sequence, respectively. It is thought that bacterial proteins harboring structural domains similar to eukaryotic transglutaminases can function as proteases or cross-linking enzymes, although presently it is unclear if DufA or DufC has any enzymatic activity (44). A section of LwpA modeled to a concanavalin A (ConA)-like lectin with 100% confidence for 30% of the protein.…”

A shotgun antisense approach to the identification of novel essential genes in Pseudomonas aeruginosa

“…For each hit, the orthologue proteins, extracted from the Ortholuge DB [30], that were found in DEG are indicated with the abbreviation of the harboring bacterial species. Bacterial species: Ab ( Acinetobacter baylyi ), Bs ( Bacillus subtilis ), Bt ( Bacteroides thetaiotaomicron ), Cc ( Caulobacter crescentus ), Ec ( Escherichia coli ), Fn ( Francisella novicida ), Hi ( Haemophilus influenzae ), Hp ( Helicobacter pylori) , Mt ( Mycobacterium tuberculosis ), Mp ( Mycoplasma pulmonis ), Mg ( Mycoplasma genitalium ), Pg ( Porphyromonas gingivalis ), Pa ( Pseudomonas aeruginosa ), Se ( Salmonella enterica ), St ( Salmonella typhimurium ), Sp ( Streptococcus pneumoniae ), Ss ( Streptococcus sanguinis ), Sa (S taphylococcus aureus ), Vc ( Vibrio cholerae ). d Previous reports [24-26] did not mention growth defects associated to deletion of popD gene. e Hit not present in DEG whose essentiality was experimentally demonstrated in P. aeruginosa [21]. …”

“…d Previous reports [ 24 - 26 ] did not mention growth defects associated to deletion of popD gene. e Hit not present in DEG whose essentiality was experimentally demonstrated in P. aeruginosa [ 21 ].…”

“…e Hit not present in DEG whose essentiality was experimentally demonstrated in P. aeruginosa [ 21 ].…”

SecDF as Part of the Sec-Translocase Facilitates Efficient Secretion of Bacillus cereus Toxins and Cell Wall-Associated Proteins

“…Two uncharacterized putative murein hydrolases (BC0991 and BC1991) were absent in the Δ secDF mutant growth medium (Table 1). Both contain a transglutaminase domain, known to facilitate intra- and interprotein crosslinks and to potentially play an important role in cell wall maturation [64]. In addition the putative cell wall binding proteins EntA (BC5239), EntB (BC2952), and EntC (BC0813), identified in the secretome of B. cereus [28], were affected at the transcriptional level (Table S5) and, in the case of EntB, also in the extracellular proteome in the secDF deletion mutant (Table S3).…”