Biomedical subjects
Publications and source records attributed to U N Streips.
Using two-dimensional polyacrylamide gel electrophoresis, the GroEL homologue of Bacillus subtilis was shown to be induced upon infection with phi 105clz, a clear plaque mutant of the temperate bacteriophage phi 105. Western blotting of one dimensional polyacrylamide gels also showed the induction of the GroEL homologue when cells were infected with phi 105clz.
Bacillus subtilis strain BUL786, a transposon-generated mutant, lacks a 97 kD protein following heat shock and has unique protein secretion properties. In this study, transduction analysis demonstrated that the properties of high temperature protease secretion, heat shock protein distribution, and loss of the 97 kD heat shock protein were all transferred as a single trait to recipient bacteria. The high temperature-specific protease released by this mutant into the supernatant was also shown to be produced during heat shock by the parental cells, but is retained intracellularly.
A stalbe L-form (Sal-1) of Bacillus subtilis was found to have retained a markedly modified chromosome-membrane association when compared to intact cells. The membrane-deoxyribonucleic acid complex of the L-form was similar to that of its parental strain in quantity and stability. Genetic analysis of the L-form membrane-deoxyribonucleic acid complex revealed enrichment for markers close to the replication origin, but not for internal markers, indicating preferential attachment of the origin of chromosomal replication to the membrane. These results are in close agreement with those found for the parental bacterial form. In contrast, the replication termius region was not preferentially attached to the membrane of the L-form, even though it is enriched in the bacterial form. The association of the chromosome with the membrane at the replication terminus does not appear to be necessary for cell growth and separation, but because the L-form divides aberrantly, it may be one of the factors required for normal deoxyribonucleic acid segregation and septation.
Deoxyribonucleic acid is released into the growth medium by Bacillus subtilis at the time of competence. This DNA is enriched for the genetic markers which have previously been demonstrated to be elevated in membrane-DNA preparations and more recently in cell wall-DNA complexes. Furthermore, the purA16/leu-8 relative marker enrichment varies with time, reaching its highest point at the time of maximal competence. Enrichment remains elevated for at least 60 min further in the competence regimen. Thr results suggest that certain genetic markers of the B. subtilis chromosome are preferentially more available to the external medium as the development of competence proceeds.
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The cell wall of Bacillus subtilis 168 contains protein that is refractory to removal by salts, detergents, and denaturants.
Mutagenicity tester strains of Bacillus and Salmonella were used to assay vinyl chloride in nutrient broth at a practical concentration level. Also screened without exogenous activation were seven potential metabolites of vinyl chloride in their pure forms as well as the related epichlorohydrin. Chlorooxirane, chloroacetaldehyde, chloroacetaldehyde monomer hydrate, chloroacetaldehyde dimer hydrate, chloroacetaldehyde trimer, and epichlorohydrin produced significant mutagenic acitivity in Salmonella typhimurium strains sensitive to base-pair mutation. A recombination repair deficient strain of Bacillus subtilis was inhibited in growth by these compounds, whereas excision repair deficient and wild type strains of Bacillus subtilis were relatively unaffected. On the basis of these assays a working hypothesis for the vinyl chloride carcinogenesis mechanism is proposed which involves chlorooxirane and chloroacetaldehyde monomer hydrate as the ultimate carcinogenic metabolites of vinyl chloride.
Several procedures were used in an attempt to prepare clean cell walls from Bacillus subtilis. The results indicate that protein and nucleic acids are tightly bound to the walls. The cleanest wall preparations were found following trichloroacetic acid extraction at 60 degrees or by extraction with 0.1N NaOH under a nitrogen atmosphere for 10 hrs. Protein denaturants, such as sodium dodecyl sulfate and concentrated guanidine hydrochloride were relatively ineffective in removing proteins and nucleic acids from the cell walls. Cell wall-bound DNA was biologically active in transformation assays.
Hydrolysis of the cell wall of Bacillus subtilis 168 by autolysins or lysozyme resulted in the exposure of glucosylated teichoic acid molecules as evidenced by increased precipitation of [14C] concanavalin A. The number of concanavalin A-reactive sites increased significantly after only limited enzymatic digestion of the walls. Quantitative analyses of [14C] concanavalin A-treated wall or wall hydrolysate complexes indicate that approximately one-half of the teichoic acid molecules are surface-exposed, whereas the remainder are probably embedded within the peptidoglycan matrix. Treatment of the cell walls with sodium dodecyl sulfate or Triton X-100 did not result in new concanavalin A-reactive sites. Partial autolysis diminished the ability of the cell walls to adsorb bacteriophage phi25. Fluorescein-labeled concanavalin A bound intensely over the entire surface of growing B. subtilis 168 cells, suggesting that teichoic acid molecules are located on the total solvent-exposed surface area of the bacteria.
Cell wall and membrane teichoic acids from several bacteria formed soluble complexes with polysaccharides and bovine plasma in alkyl alcohol solutions. Polysaccharides which contain different monomeric units and anomeric configurations complexed with the teichoic acids, suggesting that the interaction is relatively nonspecific. Teichoic acids complexed glycogen or bovine plasma albumin in 50 to 97% ethanol solutions. The macromolecular association between teichoic acids and polysaccharides or proteins was independent of teichoic acid size over a threefold molecular weight range. Glycerol phosphates or an acid hydrolysate of teichoic acid would not complex to either glycogen or bovine plasma albumin in ethanol. The optimal interaction between glycogen and the Bacillus subtilis lipoteichoic acid occurred between pH 4.5 and 8.2. The ability of teichoic acids to bind polysaccharides and proteins in moderate dielectric constant solvents suggests that these polymers may serve as complexing agents for hydrophilic molecules found in membranes.