Surveying biotransformations with à la carte genetic traps: translating dehydrochlorination of lindane (gamma-hexachlorocyclohexane) into lacZ-based phenotypes.

The ability of the product of a desired reaction to activate a bacterial transcriptional regulator was exploited to develop genetic traps that render the catalytic activity born by a DNA clone into a selectable/scorable phenotype. We established this strategy with a system to expose the activity of dehydrochlorinases acting upon gamma-hexachlorocyclohexane (gamma-HCH or lindane). To this end, the effector-binding protein, XylR, was evolved by gene shuffling plus mutagenic polymerase chain reaction to be optimally responsive to the major product of gamma-HCH dehydrochlorination, 1,2,4-trichlorobenzene (TCB).

On the natural selection and evolution of the aerobic phototrophic bacteria.

This contribution gives a brief survey of the short history since the discovery of the aerobic phototrophic bacteria to focus on a general evolutionary scenario. Most of the citations are of reviews that have covered the earlier literature and to which the reader is directed at appropriate places in the following text. The data summarized in these reviews are supplemented with information from recent or otherwise key primary publications in order to support a synthesis that addresses vexing questions about bacteria containing photosynthetic pigment-protein complexes, but which are incapable of growth with light as the sole, or even the major source of energy.

Effects of Precise Deletions in Rhodobacter sphaeroides Reaction Center Genes on Steady-state Levels of Reaction Center Proteins: A Revised Model for Reaction Center Assembly.

Possible interactions between photosynthetic reaction center (RC) proteins that protect these membrane proteins from proteolytic digestion in RC complex assembly were evaluated by use of translationally in-frame (nonpolar) RC gene-specific deletions. The RC H, RC M and RC L proteins were produced from plasmids, either alone or in concert with one or both of the others, in a strain of Rhodobacter sphaeroides that contained chromosomal deletions of all three RC genes. The steady-state amounts of these proteins in cell membrane and soluble fractions were assessed in western blots.

The PufX protein of Rhodobacter capsulatus affects the properties of bacteriochlorophyll a and carotenoid pigments of light-harvesting complex 1.

A pufX gene deletion in the purple bacterium Rhodobacter capsulatus causes a severe photosynthetic defect and increases core light-harvesting complex (LH1) protein and bacteriochlorophyll a (BChl) levels. It was suggested that PufX interrupts the LH1 alpha/beta ring around the reaction centre, allowing quinone/quinol exchange. However, naturally PufX(-) purple bacteria grow photosynthetically with an uninterrupted LH1.

Catabolism of benzoate and phthalate in Rhodococcus sp. strain RHA1: redundancies and convergence.

Genomic and proteomic approaches were used to investigate phthalate and benzoate catabolism in Rhodococcus sp. strain RHA1, a polychlorinated biphenyl-degrading actinomycete. Sequence analyses identified genes involved in the catabolism of benzoate (ben) and phthalate (pad), the uptake of phthalate (pat), and two branches of the beta-ketoadipate pathway (catRABC and pcaJIHGBLFR).

The puhE gene of Rhodobacter capsulatus is needed for optimal transition from aerobic to photosynthetic growth and encodes a putative negative modulator of bacteriochlorophyll production.

A conserved orf of previously unknown function (herein designated as puhE) is located 3' of the reaction centre H (puhA) gene in purple photosynthetic bacteria, in the order puhABCE in Rhodobacter capsulatus. Disruptions of R. capsulatus puhE resulted in a long lag in the growth of photosynthetic cultures inoculated with cells grown under high aeration, and increased the level of the peripheral antenna, light-harvesting complex 2 (LH2).

The PuhB protein of Rhodobacter capsulatus functions in photosynthetic reaction center assembly with a secondary effect on light-harvesting complex 1.

The core of the photosynthetic apparatus of purple photosynthetic bacteria such as Rhodobacter capsulatus consists of a reaction center (RC) intimately associated with light-harvesting complex 1 (LH1) and the PufX polypeptide. The abundance of the RC and LH1 components was previously shown to depend on the product of the puhB gene (formerly known as orf214). We report here that disruption of puhB diminishes RC assembly, with an indirect effect on LH1 assembly, and reduces the amount of PufX.

Evolutionarily divergent extradiol dioxygenases possess higher specificities for polychlorinated biphenyl metabolites.

The reactivities of four evolutionarily divergent extradiol dioxygenases towards mono-, di-, and trichlorinated (triCl) 2,3-dihydroxybiphenyls (DHBs) were investigated: 2,3-dihydroxybiphenyl dioxygenase (EC 1.13.11.

Cyclic dipeptides exhibit synergistic, broad spectrum antimicrobial effects and have anti-mutagenic properties.

Cyclic dipeptides are known to have antiviral, antibiotic and antitumour properties. The aim of this study was to determine the combined effects of cyclo(L-leucyl-L-prolyl) and cyclo(L-phenylalanyl-L-prolyl) on the growth of vancomycin-resistant enterococci (VRE) and pathogenic yeasts, as well as determining their anti-mutagenic effects. This drug combination was especially effective against five VRE strains: Enterococcus faecium (K-99-38), E.

Steady-state kinetics and inhibition of anaerobically purified human homogentisate 1,2-dioxygenase.

HGO (homogentisate 1,2-dioxygenase; EC 1.13.11.

Stability of the bacterial community in a pulp mill effluent treatment system during normal operation and a system shutdown.

Currently, very little is known about the normal dynamics of microbial populations in wastewater treatment systems and the relationship between population dynamics and functional stability of treatment systems. We monitored the bacterial community in an oxygen activated sludge system at a pulp and paper mill during a 55-day period that included normal operation as well as an 11-day shutdown of the system and the subsequent start-up. Ribosomal intergenic spacer (RIS) length polymorphism fingerprints were very similar (57-88% similar) throughout the study period.

Cationic antimicrobial peptides activate a two-component regulatory system, PmrA-PmrB, that regulates resistance to polymyxin B and cationic antimicrobial peptides in Pseudomonas aeruginosa.

The two-component regulatory system PhoP-PhoQ of Pseudomonas aeruginosa regulates resistance to cationic antimicrobial peptides, polymyxin B and aminoglycosides in response to low Mg2+ conditions. We have identified a second two-component regulatory system, PmrA-PmrB, that regulates resistance to polymyxin B and cationic antimicrobial peptides. This system responds to limiting Mg2+, and is affected by a phoQ, but not a phoP mutation.

A conserved region within the Bordetella pertussis autotransporter BrkA is necessary for folding of its passenger domain.

Autotransporter secretion represents a unique mechanism that Gram-negative bacteria employ to deliver proteins to their cell surface. BrkA is a Bordetella pertussis autotransporter protein that mediates serum resistance and contributes to adherence of the bacterium to host cells. BrkA is a 103 kDa protein that is cleaved to form a 73 kDa alpha-domain and a 30 kDa beta domain.

Evidence for a role for the Dictyostelium Rap1 in cell viability and the response to osmotic stress.

The Dictyostelium genome contains a single rapA gene, which encodes a Rap1 monomeric G protein. As attempts at generating rapA-null Dictyostelium cells had been unsuccessful, expression of antisense RNA from the rapA gene under control of the folate repressible discoidin promoter was used to reduce cellular levels of the Rap1 protein. As Rap1 levels gradually decreased following antisense rapA RNA induction, growth rate and cell viability also decreased, a result consistent with the idea that rapA is an essential gene.

Role of membranes in the activities of antimicrobial cationic peptides.

Cationic amphiphilic peptides that are found throughout nature have very broad-spectrum activities against microbes. The initial sites of interaction are with microbial membranes. Although dogma suggests that their lethal action involves disruption of the cytoplasmic membranes, a number of cationic peptides can traverse intact membranes to interact with internal targets.

Identification of lipopolysaccharide O antigen synthesis genes required for attachment of the S-layer of Caulobacter crescentus.

The outer surface of Caulobacter crescentus consists of a two-dimensional crystalline protein lattice layer (S-layer). A fraction of the LPS has an O antigen polymer attached to the core to form a 'smooth' LPS (S-LPS), which is required for attachment of the protein S-layer to the outer-membrane surface. A method to screen for strains defective in LPS production, based on loss of S-layer attachment, was developed and applied to libraries of transposon-generated mutants.

The role of cationic antimicrobial peptides in innate host defences.

Cationic antimicrobial peptides are found in all living species. A single animal can contain >24 different antimicrobial peptides, which fall into four structural classes. These peptides are produced in large quantities at sites of infection and/or inflammation and can have broad-spectrum antibacterial, antifungal, antiviral, antiprotozoan and antisepsis properties.