Purification and characterisation of a novel 34,000-Mr cell-associated proteinase from the dermatophyte Trichophyton rubrum

Abstract A novel cell-associated proteinase was purified to homogeneity from cytoplasmic antigen preparations of Trichophyton rubrum by sequential isoelectric focusing and gel filtration chromatography. The enzyme exhibited relative molecular masses of 34,000- M _r (non-reduced sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE)), 15,000- M _r (reduced SDS-PAGE) and 37,000- M _r (substrate SDS-PAGE). It had a pH optimum of 7.5 and a p I of 4.5. The proteinase exhibited broad substrate specificity and it was strongly inhibited by the serine proteinase inhibitors phenylmethylsulfonyl fluoride and chymostatin. The N-terminal amino acid sequence of the 34,000- M _r proteinase shared 50% homology with the deduced amino acid sequence of a Coccidioides immitis wall-associated chymotrypsin-type serine proteinase. This is the first cell-associated proteinase to be purified and characterised from T. rubrum and it would appear to be related to the chymotrypsin-type serine proteinases, a class of enzymes that have rarely been isolated from fungi. The function of the proteinase remains speculative although it may play a role in the development and subsequent proliferation of the fungus in vivo.     

Purification and properties of a novel polyol dehydrogenase of bacterial origin

Abstract A bacterium, as yet unidentified, has been isolated from floor dust by direct selection on minimal agar using l -glucitol ( d -gulitol) as the sole carbon energy source. The bacterium possesses a constitutive enzyme which catalyzes the reaction: l -glucitol + NAD⁺→ d -sorbose + NADH + H⁺. A new species of enzyme has been induced by l -arabinitol or ribitol, but not l - or d -glucitol, and the induction is only partially counteracted by the glucose-repression effect. The constitutive enzyme was purified by fractionation on Sephadex G-200 gel and chromatography on DEAE Biogel A. The enzyme required NAD⁺, but not NADP⁺, as a cofactor. It oxidizes also ribitol, xylitol and l -arabinitol, but not d -arabinitol, lactitol or a variety of other commercially available alditols. The enzyme is not inhibited by 10 mM sodium azide but is totally inhibited by 0.1 mM potassium ferricyanide.     

Activation of potassium channels during metabolite detoxification in Escherichia coli

In bacteria the detoxification of compounds as diverse as methylglyoxal and chlorodinitrobenzene proceeds through the formation of a glutathione adduct. In the Gram-negative bacteria, e.g. Escherichia coli, such glutathione adducts activate one, or both, of a pair of potassium efflux systems KefB and KefC. These systems share many of the properties of cation-translocating channels in eukaryotes. The activity of these systems has been found to be present in a range of Gram-negative bacteria, but not in the glutathione-deficient species of Gram-positive organisms. The conservation of the activity of these systems in a diverse range of organisms suggested a physiological role for these systems. Here we demonstrate that in E. coli cells activation of the KefB efflux system is essential for the survival of exposure to methylglyoxal. Methylglyoxal can be added to the growth medium or its synthesis can be stimulated in the cytoplasm. Under both sets of conditions survival is aided by the activity of KefB. Inhibition of KefB activity by the addition of 10 mM potassium to the growth medium stimulates methylglyoxal-induced cell death. This establishes an essential physiological function for the KefB system.     

The Energetic Determination, Spatial Dispersion and Density Dependence of Myrmeleon Ant Lion Pits in Las Cruces, Costa Rica

The amount of space used by an organism is energetically determined. We utilized a population of ant lion larvae in Costa Rica to test allometric theories concerning the use of space by organisms and how different densities of individuals affect the use of space. The area of ant lion trapping pits scaled with mass to the three-quarters power, supporting allometric theory for sessile organisms. Our analyses also show that larger ant lion larvae show spatial repulsion and facultative density dependent pit-building strategies.     

First record of Aulacidae (Hymenoptera: Evanioidea) from New Caledonia with descriptions of three new species of Aulacus Jurine

Abstract The parasitoid wasp family Aulacidae is recorded for the first time from New Caledonia, and three new species are described: Aulacus burwelli, A. coracinus , and A. emineo . Although recorded from just a few specimens, these species appear to be restricted to the moist evergreen rainforest areas of the main island of Grande Terre. They appear unrelated to Australian members of the family, based on a number of characters associated with the forewing venation and hind coxal morphology, as well as size and colour. A key to distinguish the species is presented, as well as information on their distribution and possible relationships.     

Additional hox clusters in the zebrafish: divergent expression patterns belie equivalent activities of duplicate hoxB5 genes

SUMMARY The evolution of metazoan body plans has involved changes to the Hox genes, which are involved in patterning the body axis and display striking evolutionary conservation of structure and expression. Invertebrates contain a single Hox cluster whereas tetrapods possess four clusters. The zebrafish has seven unlinked hox clusters, a finding that is difficult to reconcile with the notion that genomic complexity, reflected by Hox cluster number, and morphological complexity are causally linked, as the body plan of the zebrafish is not obviously more complex than that of the mouse or human. Why have the additional hox genes in zebrafish been conserved? To address the role of these additional zebrafish hox genes, we have examined the duplicate hoxB5 genes, hoxB5a, and hoxB5b. Conservation of gene duplicates can occur when one gene acquires a new function (neofunctionalization), or when the ancestral function is divided between the two duplicates (subfunctionalization). hoxB5a and hoxB5b are expressed in distinct domains, and their combined expression domain is strikingly similar to that of single Hoxb5 genes in other species. The biochemical functions encoded by the two genes were studied by overexpression, which resulted in identical developmental defects in the anterior hindbrain and cranial neural crest, suggesting strongly that hoxB5a and hoxB5b have equivalent biochemical properties with respect to early development. From these studies, we conclude that conservation of hoxB5a and hoxB5b is likely the result of division of the ancestral Hoxb5 function between the two genes, without significant changes in biochemical activity. These results suggest a resolution to the conundrum of the extra hox genes and clusters in the zebrafish, since if any of the additional hox genes in the zebrafish are similarly subfunctionalized, they are unlikely to supply novel genetic functions. Thus, the morphological complexity potentially conferred by the majority of additional zebrafish hox clusters may not be substantially greater than that conferred by the four tetrapod clusters.     

The sym plasmid pRP2JI and at least two other loci of Rhizobium leguminosarum biovar phaseoli can confer resistance to infection by the virulent bacteriophage RL38

Abstract The virulent Rhizobium bacteriophage RL38 did not form plaques on R.leguminosarum by phaseoli but did so at high efficiency on a derivative of that strain lacking its symbiotic plasmid pRP2JI. Other strains with large deletions in pRP2JI which removed many nod and nif genes retained resistance to RL38, showing that the gene which confers phage resistance lies elsewhere on the plasmid. Although the wild-type strain of R. leguminosarum bv. phaseoli failed to plate RL38, it was possible to transduce chromosomal markers into this strain, indicating that the 'block' was not at an early stage in the infection process. Two different recombinant plasmids obtained from a clone bank of genomic DNA of R. leguminosarum bv. phaseoli , which appeared to have no DNA in common, both conferred resistance to RL38. Surprisingly, the DNA cloned in each of these plasmids did not originate from pRP2JI. Therefore, several different loci both on the Sym plasmid and elsewhere on the bacterial genome can be involved in conferring resistance to this bacteriophage.