Aspartate transcarbamoylase (ATCase) of Streptomyces was characterized and its interaction with other pyrimidine enzymes explored.

Aspartate transcarbamoylase (ATCase) catalyzes the first committed step in pyrimidine biosynthesis. Bacterial ATCases are divided into three classes, A, B and C. Class A ATCases are largest at 450-500, are. dodecamers and represented by Pseudomonas ATCase. The overlapping pyrBC' genes encode the Pseudomonases ATCase, which is active only as a 480 kDa dodecamer and requires an inactive pyrC'-encoded DHOase for ATCase activity. ATCase has been studied in two non-pathogenic members of Mycobacterium, M. smegmatis and M. phlei. Their ATCases are dodecamers of molecular weight 480 kDa, composed of six PyrB and six PyrC polypeptides. Unlike the Pseudomonas ATCase, the PyrC polypeptide in these mycobacteria encodes an active DHOase. Moreover, the ATCase: DHOase complex in M. smegmatis is active both as the native 480 kDa and as a 390 kDa complex. The latter lacks two PyrC polypeptides yet retains ATCase activity. The ATCase from M. phlei is similar, except that it is active as the native 480 kDa form but also as 450,410 and 380 kDa forms. These complexes lack one, two, and three PyrC polypeptides, respectively. By contrast,.ATCases from pathogenic mycobacteria are active only at 480 kDa. Mycobacterial ATCases contain active DHOases and accordingly. are placed in class A1 . …

Salvage of pyrimidine nucleosides and bases by S. griseus and the regulation of aspartate transcarbamoylase (ATCase) were studied. The velocity-substrate curve for S. griseus ATCase was hyperbolic for both aspartate and carbamoylphosphate. The enzyme activity was diminished in the presence of ATP, CTP, or UTP. The synthesis of ATCase was repressed in cells grown in the presence of exogenous uracil. The specific activity of cells grown with uracil was 43 percent of that for cells grown in minimal medium only. Maximal ATCase and dihydroorotase activities were found in the same column fraction after size-exclusion chromatography, suggesting that both activities could reside in the same polypeptide. The pyrimidine salvage enzymes cytosine deaminase and uridine phosphorylase were identified in S. griseus using HPLC reversed-phase chromatography.

Bacterial aspartate transcarbamoylases (ATCase's) are divided into three classes that correspond to taxonomic relationships within the bacteria. The opportunistic pathogen Moraxeila catarrhalis has undergone several reclassifications based on traditional microbiological criteria. The previously uncharacterized ATCase from M. catarrhalis was purified to homogeneity and its chemical properties characterized. The ATCase from M. catarrhalis is a class C ATCase with an apparent molecular mass of 480-520 kDa. The M. catarrhalis ATCase is a dodecomer composed of six 35 kDa polypeptides and six 45 kDa polypeptides. The enzyme has an unusually high pH optimum of greater than pH 10. The enzyme exhibited hyperbolic kinetic with a Km for aspartate of 2 mM. A single, separate 78 kDa dihydroorotase from M. catarrhalis was identified and it was not associated with ATCase. These data support the reclassification of M. catarrhalis out of the Neisseriaceae family.

Cardiorespiratory responses to incremental dynamic exercise were assessed across four different levels of lower-body positive pressure (LBPP) and, as a separate study, during constant load (i.e constant work rate) exercise below and above each subject's ventilatory threshold (VT), both with and without 45 torr of LBPP.

Cassette systems for creating intergeneric hybrid ATCases were constructed. An MluI restriction enzyme site was introduced at the carbamoylphosphate binding site within the pyrB genes of both Pseudomonas putida and Escherichia coli. Two hybrids, E. coli pyrB polar domain fused with P. putida pyrB equatorial domain and P. putida pyrB polar domain fused with E. coli pyrB equatorial domain, are possible. The intergeneric E. coli-P. putida hybrid pyrB gene was constructed and found to encode an active ATCase which complemented an E. coli Pyr- strain. These hybrids are useful for kinetic and expression studies of ATCase in E. coli.

Olestra is, a sucrose polyester, a noncaloric fat substitute, made from sucrose and several fatty acid esters. It has been approved by the FDA as a food additive used in preparing low-fat deep-frying foods such as savory snacks. Available literature on olestra was evaluated that had both positive and negative connotations. Clinical trials in numerous species of animals including humans were conducted to determine if olestra would affect the utilization and absorption of macro- and micronutrients; the effects of olestra on growth, reproduction, or its toxicity were also examined. The roles of olestra as a fat substitute, how it could effect on humans and the environment, and the potential impacts from its use in large amounts were assessed. Olestra can be removed from the environment by aerobic bacteria and fungi which may be isolated from activated sludge and soils.

The effects of single injections and daily oral administration of ethanol on plasma levels of ACTH, beta endorphin, and corticosterone in response to cold stress were examined. The long-term experimental animals were given 0.25 ml of 28% ethanol or water orally once a day, five days a week, for fourteen months. Plasma levels of ACTH, beta endorphin, and corticosterone were lower in alcohol-treated rats as compared with water-treated rats when exposed to cold stress. The effects of a single injection of ethanol significantly elevated plasma levels of all three hormones. Mortality in sham-treated males was higher than ethanol-treated.

Concern over sediment toxicity has increased the need for toxicity test information with organisms that inhabit sediments. Oligochaetes are exposed to toxicants through feeding and direct body contact with aquatic sediments. Chronic testing with oligochaetes has historically focused on tubificids with test lengths of one year or more to encompass several generations. Most naidid oligochaetes have generation times of three to seven days and could provide chronic information in a matter of weeks. The cosmopolitan distributed naidid, Pristina leidyi, was evaluated for use as a toxicity test organism. Results of research conducted includes culture methods, effects of temperature on reproduction, growth rates in a reference sediment, acute toxicity tests, and chronic toxicity tests.

Acclimation temperature had a significant effect on the lethality of copper to fathead minnows in replicated 96-hour lethality tests. Lowest median lethal copper concentrations (LC50) were recorded at 12 and 22°C, with LC50s at 5 and 32°C at least 140 μg/1 higher. This research found LC50 copper concentrations in the 300 to 500 μg/1 range and a polynomial relationship between LC50s and acclimation temperature. Following a 24-hour exposure to three sublethal concentrations of copper, critical thermal maxima (CTMax) were tested in minnows acclimated to four temperatures. Sublethal exposure to copper significantly decreased the CTMaxs relative to controls at three of the four temperatures. Control CTMaxs ranged from 28.6 to 40.4°C and increased 0.46°C for each 1°C increase in acclimation temperature.

The biosynthesis of pyrimidines in Pseudomonas putida was investigated. In this study, pyrimidine requiring mutants were isolated by conventional mutagenesis and enrichment. The strains required exogenously supplied pyrimidines for growth and were found by enzyme assays to be deficient for the product of the pyrB gene encoding the enzyme aspartate transcarbamoylase. None of the intermediates of the pathway could supply the auxotrophic requirement of the strain; only preformed pyrimidines, metabolized via salvage pathways could suffice. Pyrimidine limitation in the mutant caused a slight but significant fifty per cent increase in expression of all the de novo biosynthetic enzymes. Pyrimidine starvation's effect on nucleotide pool levels was examined in the mutant and caused a marked swelling of the purine nucleotide pools.

The enzyme that catalyzes the committed step in pyrimidine biosynthesis, aspartate transcarbamoylase, has been compared in selected endospore-forming organisms and in morphologically similar control organisms. The ATCases and pyrimidine salvage from Sporosarcina ureae, Sporolactobacillus inulinus, Lactobacillus fermentum, and Micrococcus luteus were compared to those of Bacillus subtilis. While the ATCases from Sporosarcina ureae, Sporolactobacillus inulinus, and L. fermentum were found to exhibit characteristics to that of Bacillus with respect to molecular weight and kinetics, M. luteus ATCase was larger at approximately 480 kDa. Furthermore, pyrimidine salvage in Sporosarcina ureae and M. luteus was identical to those of B. subtilis, while pyrimidine salvage of Sporolactobacillus inulinus and L. fermentum resembled that of the pseudomonads.

Research estimated toxicity of refuse-derived fuel fly ash (RDF-FA) on two earthworms species, Lumbricus terrestris and Eisenia foetida. Specific objectives were to: (1) Compare their 14-day LC50s under light and dark conditions; (2) separate toxicity due to osmotic, pH and physical factors from that of heavy metal contaminants; (3) compare relative differences of artificial soil and commercial soil as exposure media for evaluating toxicity to earthworms. The 14-d LC50s for L. terrestris in dark and light were 57.0 and 48.34 % RDF-FA, and 59.25 and 41.00 % RDF-FA for E. foetida using artificial soil. All of the toxicity resulted from heavy metals within the RDF-FA. Using L. terrestris, the LC50s for artificial soil and commercial soil were 52.30 and 64.34%.

Pseudomonas fluorescens does not appear to regulate the enzymes of de novo pyrimidine biosynthesis at the level of gene expression. Little or no apparent repression of pyr gene expression is observed upon addition of exogenous pyrimidines to the growth medium. The Escherichia coli pyrBI genes for aspartate transcarbamoylase (ATCase) were sized down and cloned into the broad host range plasmid, pKT230. Upon introduction into a P.fluorescenspyrB mutant strain, ATCase showed repression in response to exogenously fed pyrimidine compounds. Thus, it was possible to bring about changes in pyrimidine nucleotide pool levels and in transcriptional regulation of gene expression at the same time.

Recent studies suggest that synthesis of the Colony-stimulating factor (CSF) is a well regulated process. However, the molecular mechanisms of the signal transduction of the various inducers of CSF such as monokines and lymphokines are not well understood. Using Interleukin 1 (IL-1) stimulation of CSF-1 in the MIA PaCa-2 cell line as a model system, the involvement of G-protein has been studied. The IL-1 induction of CSF-1 synthesis can be inhibited by both Pertussis toxin and Cholera toxin, which are known to modify the Gᵢ and Gₛ proteins respectively, thus activating adenylate cyclase to release more cAMP. The toxin inactivation can be prevented by inhibitors of the ADP-ribosylation such as, benzamide and MBAMG. Addition of dibutyryl-cAMP inhibits the IL-1 induced CSF production. Both Theophylline and Forskolin which increase cAMP by inhibiting phosphodiesterase and stimulating adenylate cyclase respectively, also inhibit CSF-1 production. Results from these studies have shown that cAMP level inversely regulates the biosynthesis of CSF-1. Preincubation of MIA PaCa-2 cells with IL-1 and 5'- guanylylimidodiphosphate (GppNHp) prevents the inhibitory effect of pertussis toxin on CSF-1 production. These data are consistent with the hypothesis that IL-1 binds to its receptor and couples to Gᵢ∝ resulting in the inhibition of adenylate …

The immunomodulatory effects of Trichinella spiralis or Trichinella pseudospiralis soluble antigen extracts were examined in an effort to characterize the differences in immune responses seen during these Trichinella infections. The newborn larvae extracts of either parasite exhibited similar potency for stimulating macrophage PGE production; however, the muscle larvae extracts of T. pseudospiralis stimulated greater levels of PGE than did the muscle larvae extracts of T. spiralis. These data clearly indicate that Trichinella antigens possess immunomodulatory capabilities.

Ancient peoples did not distinguish between philosophy, religion, and science. Scientific truth did not exist apart from divine truth. Any new idea, finding, or theory was assimilated into a monolithic mythological structure. This is one of the causes of the underestimation of ancient science: it is always packaged in a myth - the method of preserving information in an oral culture. The mythological medium allowed the preservation and dissemination of hard-won, empirical, scientific knowledge through generations of preliterate peoples. The context for mythological memorization, or simply tradition, needed to be easily and naturally acquired. The ideal context was the anthropomorphic context, the ontogenic context. This is the Grand Allegory - the anthropomorphization of information. Biomyths are essentially biological texts allegorized in esoteric language.

Six reagent waters, eleven diets, and two culture sizes were evaluated for culturing C. dubia. Different filtration techniques were used to prepare the reagent waters. The eleven diets were comprised of two algae augmented with eight supplements. Reproduction and growth were assessed to discern differences among C. dubia raised in mass cultures and cultured in individual cups, during which, bacterial population densities, lipid, protein, and carbohydrate concentrations of the diets were measured. Results showed that a glass-distilled, carbon filtered, deionized reagent water and a Selenastrum capricornutum- Cerophyl® diet were optimum for culturing. Mass culturing supported the highest reproduction and growth, while no correlation was found between nutritional measurements and production.

Wetland wastewater treatment offers low-cost, energy efficient alternatives to conventional wastewater technologies. In this study, an artificial wetland was constructed at the City of Denton, Texas Pecan Creek Water Reclamation Plant to facilitate diazinon removal from treated effluent.

Growth of mussels under laboratory conditions was examined under various food regimes in different water types and temperatures. Growth was less than would be useful as an indicator and comparisons with field exposures were of minimal value. The effects of organophosphates on bivalves were examined via toxicity tests, tissue concentration, and by controlling exposure through the use of physical constraints. Upper temperature tolerance of both bivalve species was examined with respect to different acclimation temperatures and organophosphate exposures. Deviations from control exposures occurred at some temperatures. Copper effectively lowered the mean heat coma temperatures of C. fluminea at some concentrations, however, chlorine exposures did not alter heat coma temperature.