摘要:

The steady-state levels of zinc inCandida utilisyeast grown in continuous culture under conditions of zinc limitation are <1 nmol Zn2+/mg dry weight of cells. Unlike carbon-limited cells, zinc-depleted cells from a zinc-limited chemostat possess the capacity to accumulate and store zinc at levels far in excess of the steady-state level of 4 nmol/mg dry biomass observed in carbon-limited chemostat cultures. Zinc uptake is energy-dependent and apparently unidirectional since accumulated65Zn neither exits from preloaded cells nor exchanges with cold Zn2+. The transport system exhibits a high affinity for Zn2+(Km = 0.36 μM) with aVmaxof 2.2 nmol per minute per milligram dry weight of cells. Growth during the period of the uptake assay is responsible for the apparent plateau level of 35 nmol Zn2+/mg dry weight of cells achieved after 20–30 min in the presence of65Zn at pH 4.5 and 30 °C. Inhibition of growth during the uptake assay by cycloheximide results in a biphasic linear pattern of zinc accumulation where the cellular zinc is about 60 nmol/mg dry weight after 1 h. The enhanced level of accumulated zinc is not inhibitory to growth. Zinc-depletedC.utiliscontains elevated amounts of polyphosphate and this anionic polymer must be considered a possible zinc-sequestering and storage agent. Present experimental evidence does not allow discrimination between possible regulation of zinc homeostasis either by inhibition of zinc efflux through control of the membrane carrier or by control of the synthesis of a cytoplasmic zinc-sequestering macromolecule.

ISSN:0008-4166    

DOI:10.1139/m80-011

出版商:NRC Research Press    

年代:1980

数据来源: NRC

摘要:

Exocellular protease production was examined in two separate strains ofPseudomonas aeruginosa, one a clinical isolate and the other a laboratory strain. Both strains produced two separate proteases (proteases 1 and 2) which were indistinguishable from one strain to the other. The two proteases were purified by a two-step procedure of gel filtration chromatography followed by ion-exchange chromatography. Proteases 1 and 2 were shown to be distinct serologically and unrelated by physicochemical parameters examined. Protease 1 was the major exocellular protein produced and contributed about 95% of the total protease activity of the culture. It was estimated to have a molecular weight of 34 850 and was also shown to contain 10% glucosamine by weight. Protease 2, in contrast, had an estimated molecular weight of 52750 and contained no detectable carbohydrate. Proteases 1 and 2 were both stimulated by Ca2+, and Mg2+and inhibited by Co2+Zn2+, and 1,10-o-phenanthroline. Protease 1 was also inhibited by EDTA. In addition to protease activity, both proteases 1 and 2 demonstrated elastase activity as well as a limited collagenase activity. Specificity of the two proteases against synthetic peptides was, however, quite different. Protease 1, but not protease 2, showed a preference for peptide bonds in which the amino group was contributed by an amino acid with a hydrophobic R group.

ISSN:0008-4166    

DOI:10.1139/m80-012

出版商:NRC Research Press    

年代:1980

数据来源: NRC

摘要:

The enzymatically active form of protease 1, the major exocellular protein produced byPseudomonas aeruginosastrain 34362, has been shown to exist exclusively exocellularly with no significant cell-associated activity. However, the presence of a cell-associated, enzymatically inactive protein which is serologically cross-reactive with, and convertible to, active enzyme has been demonstrated. One method of conversion of "precursor" to active enzyme is via limited proteolysis. Two assay systems for precursor were developed, one a radioimmune assay, and the other a proteolytic activation procedure. Localization studies suggest that the cell-associated precursor resides primarily in the envelope (periplasmic) fraction, and that the association while more tenacious than classical periplasmic enzymes is still an ionic rather than a covalent one. Kinetics of production studies showed the precursor to be synthesized early in the growth cycle and to accumulate prior to the rapid release of the active enzyme. Molecular weight studies showed only slight changes produced upon activation.

ISSN:0008-4166    

DOI:10.1139/m80-013

出版商:NRC Research Press    

年代:1980

数据来源: NRC

摘要:

Two plasmids, pLK1 and pLK2, were derived from pIP218, anin vivorecombinant of plasmid F and the drug-resistance plasmid pIP176 (Cmr, Smr, Sur, Tcr). Of these two plasmids, pLK1 is 70 Mdaltons and carries the Tc-resistance determinant in a 7-Mdalton transposition element; pL K2 is 125 Mdaltons and carries Cm-, Sm- and Su-resistance determinants.The plasmid pLK1 resulted as a Tc-resistance segregant of pIP218 during its coexistence with another plasmid, Co1E1-araC 101, and pLK2 (125 Mdaltons) as a CmrSmrSursegregant during the conjugal transfer of pIP218. Both plasmids belong to the F1-incompatibility group, surface-exclude each other and Flac, and are derepressed for transfer. Incompatibility studies also indicated the preferential maintenance of pLK2 in hosts carrying either pLK2 and pLK1, or pLK2 and F'lac. An explanation of this phenomenon is provided. Furthermore, our data suggest the illegitimate recombination of the chromosomallacgenes with pLK1. In course of the incompatibility studies, thetetdeterminant was transposed from pLK1 into the chromosome, from the chromosome into thelacgenes of an Flacplasmid, and from the Flacplasmid into another site on a second Flacplasmid.

ISSN:0008-4166    

DOI:10.1139/m80-014

出版商:NRC Research Press    

年代:1980

数据来源: NRC

摘要:

The yeast-mycelial transition inCandida albicanscan be induced from yeast cells grown on minimal defined medium only in stationary phase. This study examined the inducibility of cultures in which growth was limited by the availability of the nutrients, glucose, NH4Cl, or galactose. The results showed that neither stationary phase nor cell cycle stage alone was a sufficient condition to support subsequent germ tube formation. In addition, final cell concentration alone was not a factor in inducibility. When a hundredfold decrease in growth was obtained by limiting any of the nutrients, a loss in inducibility was observed. However, the loss of inducibility differed with the limiting nutrient. Galactose, NH4Cl, and glucose-limited cultures showed respectively 15, 30, and 80% loss of inducibility. Thus the effect was associated with both carbon/energy and nitrogen-limited cells; however, glucose appeared to have a specific effect. These observations suggest that the metabolic state of the stationary phase yeast cell was an important factor in the subsequent ability to respond to conditions inducing germ tube formation.

ISSN:0008-4166    

DOI:10.1139/m80-015

出版商:NRC Research Press    

年代:1980

数据来源: NRC