Structure of the Peptidoglycan Synthase Activator LpoP in Pseudomonas aeruginosa

1. Download : Download high-res image (234KB)
2. Download : Download full-size image

     

Peptidoglycan lytic activity of the Pseudomonas aeruginosa phage phiKZ gp144 lytic transglycosylase

The gp144 endolysin gene from the Pseudomonas aeruginosa phage phiKZ was cloned and studies of gp144 expression into Escherichia coli showed host cell lysis. The gp144 protein was purified directly from the culture supernatant and from the bacterial cell pellet and showed in vitro antibacterial lytic activity against P. aeruginosa bacteria and degraded purified peptidoglycan of Gram-negative bacteria. MS analysis identified the gp144 peptidoglycan cleavage site and confirmed a lytic transglycosylase enzyme. Studies of gp144 expression in the presence of sodium azide (NaN(3)), an inhibitor of the protein export machinery, and into an E. coli MM52 secA(ts) mutant at permissive and restrictive temperatures showed that gp144 was secreted independently of the Sec system. The solution conformation of purified gp144 analyzed by circular dichroism spectroscopy was 61% in alpha-helical content, and showed a 72% decrease when interacting with dimyristoylphosphatidylglycerol (DMPG), one of the major components of bacterial membranes and less than 10% with dimyristoylphosphatidylcholine (DMPC) found in eukaryotic membranes. Membrane vesicles of DMPG anionic lipids containing calcein indicated that gp144 caused a rapid release of fluorescent calcein when interacting with synthetic membranes. These results indicated that gp144 from phiKZ is a lytic transglycosylase capable of interacting with and disorganizing bacterial membranes and has potential as an antipseudomonal in phage therapy.     

Pseudomonas aeruginosa exotoxin

Different P. aeruginosa strains have been found to differ in exotoxin synthesis. The strain isolated at the Mechnikov Research Institute for Vaccines and Sera (Moscow) and newly isolated cultures obtained from patients with the severe course of the infectious process have been found to possess the highest toxigenic activity and to synthesize exotoxins with the most complete set of pathogenically important antigens. The technological scheme for the production of stable exotoxin which can be used for the development of diagnostic, therapeutic and prophylactic preparations against Pseudomonas infections is proposed.     

Pseudomonas aeruginosa toxoid

P. aeruginosa adsorbed toxoid has been obtained. The stabilization of exotoxins and the content of proteases, hemolysin, lecithinase in their structure have been found to enhance the immunogenic potency of preparations which protect test animals from death caused by the experimental injections of toxins, homologous and heterologous to bacterial strains of different O-serogroups, into these animals. Antibodies neutralizing the lethal action of P. aeruginosa exotoxin have been detected in the blood sera of immunized animals.     

Dissociation in Pseudomonas aeruginosa

Zierdt, C. H. (National Institutes of Health, Bethesda, Md.), and P. J. Schmidt. Dissociation in Pseudomonas aeruginosa. J. Bacteriol. 87:1003-1010. 1964.-Evidence is presented that dissociation of Pseudomonas aeruginosa occurs in vivo as well as in vitro, although it is suppressed in the blood stream. Of 116 primary cultures on blood agar, 77 (66%) had more than one colony type, with a range of 2 to 6 types per culture. Dissociation was studied in 14 primary cultures during 30 serial blood agar passages. Six of these did not dissociate. Of the six, three were originally primary monocolony strains, and three were strains with two colonial types. Seven of the remaining eight cultures had more than one colony type on the primary culture plate. These eight cultures were observed to dissociate at varying rates; 25 morphological and biochemical tests failed to reveal important differences in the colonial dissociants. However, they may be differentiated by bacteriophage action. Colonial morphology in a given strain of P. aeruginosa can be correlated with its bacteriophage lytic pattern, but patterns frequently undergo drastic change during subculture of the organism. The frequently seen different colonial forms in a specific primary culture are usually related, as proven by bacteriophage typing. Phenotypic colony changes after lysogenization were observed. Mucoid colonial variants are markedly more sensitive than are the nonmucoid to streptomycin, tetracycline, and chloramphenicol.     

Pseudomonas aeruginosa melanin]

The EPR, IR, UV and visible absorption spectra of a melanin-like pigment from P. aeruginosa were studied. By the whole complex of the spectral characteristics the pigment may be classified as belonging to the melanin group. The study of the antibiotic properties of the pigment showed that it did not inhibit the growth of P. aeruginosa and E. coli. Still, it possessed some antistaphylococcal activity.     

Mesosomes in Pseudomonas aeruginosa

The use of a combination fixative-staining procedure has allowed a detailed observation of mesosomes in thin sections of Pseudomonas aeruginosa.     

The pyocins of Pseudomonas aeruginosa

Pyocins are produced by more than 90% of Pseudomonas aeruginosa strains and each strain may synthesise several pyocins. The pyocin genes are located on the P. aeruginosa chromosome and their activities are inducible by mutagenic agents such as mitomycin C. Three types of pyocins are described. (i). R-type pyocins resemble non-flexible and contractile tails of bacteriophages. They provoke a depolarisation of the cytoplasmic membrane in relation with pore formation. (ii). F-type pyocins also resemble phage tails, but with a flexible and non-contractile rod-like structure. (iii). S-type pyocins are colicin-like, protease-sensitive proteins. They are constituted of two components. The large component carries the killing activity (DNase activity for pyocins S1, S2, S3, AP41; tRNase for pyocin S4; channel-forming activity for pyocin S5). It interacts with the small component (immunity protein). The synthesis of pyocins starts when a mutagen increases the expression of the recA gene and activates the RecA protein, which cleaves the repressor PrtR, liberating the expression of the protein activator gene prtN. R and F-pyocins are derived from an ancestral gene, with similarities to the P2 phage family and the lambda phage family, respectively. The killing domains of S1, S2, AP41 pyocins show a close evolutionary relationship with E2 group colicins, S4 pyocin with colicin E5, and S5 pyocin with colicins Ia, and Ib.