Carbamoyl phosphate synthetase (CPS) catalyzes the formation of carbamoyl phosphate (CP) from MgATP, bicarbonate, and glutamine. It has three active sites, one present on the small subunit and the two phosphorylation sites present on the large subunit. These two nucleotide binding sites are homologous. Six compounds were designed to mimic the reactive intermediate species carboxy phosphate, and product cabamoyl phosphate. The apparent Ki values calculated estimated the inhibitory strengths of these compounds. These plots were also utilized in identifying the linear inhibitors, nonlinear inhibitors and partial inhibitors. Inhibition patterns were obtained with these compounds using various assay formats. Partial inhibition displayed by phosphono formate for the full biosynthetic reaction can be utilized in support of the sequential mechanism for CPS.

Phosphate is an essential but limited macronutrient for all plants. In response to limited levels of phosphate, plants have developed highly specialized developmental, biochemical, and molecular responses. To further expand our knowledge of the phosphate starvation induced signal transduction pathway in plants, the expression of the phosphate starvation inducible Purple Acid Phosphatase 1 (PAP1) gene was studied in transgenic Arabidopsis. While few components have been identified regulating gene expression under phosphate starvation conditions in plants, one cis regulatory element recognized by the MYB transcriptions factor Phosphate Starvation Response 1 (PHR1) has been identified in many phosphate starvation induced (PSI) genes. PAP1 and many other genes examined during the course of the mutant characterization contain this cis element. Using the GUS reporter gene under control of the PAP1 promoter, a mutant screen was devised for plants showing abnormal PAP1 response to phosphate nutrition. Three mutant lines were identified and subsequently characterized for the phosphate starvation-induced signal-transduction pathway in Arabidopsis. Two mutants, BT1 and BT2, both with dominant mutations, showed increased GUS staining. The mutations in BT1 and BT2 are tightly linked to the transgene and to each other, but complementation analysis suggested that they are in different genes. Characterization of these mutants indicated that the PSI genes PAP1 and At4 (in BT1 roots), and RNS1 (in BT2 leaves) have alternative or additional methods of regulation other than PHR, even though these genes all contain PHR1 binding sites. A third mutant, BT3, had a phenotype similar to the PAP1 null-mutant and did not show PAP1 phosphatase activity under normal soil-grown conditions. Characterization of BT3 indicates that PAP1, RNS1, and AtIPS1 are not exclusively regulated by PHR1. In an attempt to map the BT3 mutant in a Columbia background by crossing with Landsberg erecta (Ler), it was discovered that the Ler ecotype does not show PAP1 phosphatase activity under normal soil-grown conditions. The PAP1 phosphatase regulatory trait, named BT5, was mapped to a 15,562 bp-region area containing only two genes between the GPA1 and ER markers on Chromosome 2.

published_or_final_version / Biochemistry / Doctoral / Doctor of Philosophy

Lead and lead-tin solder were widely used in premise plumbing in North America until the 1980’s. Although this soldering practice has reduced significantly, lead release from older premise plumbing remains a concern. Utilities often use corrosion inhibitors to protect the piping in the distribution system and reduce lead release in premise plumbing. This study compares sodium silicates and phosphates as corrosion inhibitors for lead release in low alkalinity water. The study was carried out at the J.D. Kline Water Treatment Plant in Halifax, Nova Scotia, Canada. The experimental design compared sodium silicates (concentration of 18mg/L) and zinc orthophosphate (concentration of 0.8mg/L) as corrosion inhibitors at a pH of 7.3 for three disinfectant treatment groups (high chloramines (3mg/L), low chloramines (1mg/L), and chlorine (1mg/L)).

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A range of ion-selective electrodes (ISEs) for the determination of nitrate has been produced based upon rubbery membranes having covalently bound betaine salt sensor molecules. The best performing electrode contained N,N,N-triallyl leucine betaine (6.5 % m/m) covalently bound to polystyrene-block-polybutadiene-block-polystyrene (SBS) (43.5% m/m), with 2-nitrophenyIoctyl ether (2-NPOE) as solvent mediator (40 % m/m) and dicumyl peroxide (DCP) as free radical initiator (10% m/m). The Nemstian slope was -59.1 mV per decade over a linear range of 1 x 10'^-5 x 10"^ mol dm'^ nitrate, a limit of detection of 0.34 pmol dm'^ nitrate and a selectivity coefiBcient for nitrate against chloride ( ^ ° N 0 3 . , CI-) of 3.4 X IQ"^. The speed of response was less than 1 minute over the linear Nerastian range. The lifetime in the laboratory exceeded 5 months with no potentiometric drift over the linear Nemstian range. Temperafure dependency (0-25°C), pH range (2-12) and a selection of interfering anions (F', CI", B r , T, SCN, CIO4", HCO3", NO2", S04^ phthalate) were studied. A field evaluation by continuous immersion in both agricultural drainage weirs and a river were undertaken. The nitrate results obtained with the ISEs compared very favourably (R^=0.99) with those obtained with a segmented-flow instmment in a concentration range 0.47-16 ppm nitrate-N. The electrodes perfonned continuously for over 5 months in mnoff water from a field and over 2 months in river water. The ISEs did not require recalibration and no deterioration in performance or fouling of the membrane surface was observed. A preliminary investigation of a phosphate ionophore based upon a heterocyclic macrocycle was also undertaken. This work, based on previous literature, resulted in a dibasic phosphate electrode having a linear Nemstian range from 3 x lO"'' to 1 x 10"^ mol dm'^, a slope of -27 mV per activity decade and a Umit of detection of 1 x 10"^ mol dm"^ HP04^".