Sources of ammonia for mammalian urea synthesis.

The initial rate of incorporation of [15N]alanine into the 6-amino group of the adenine nucleotides in rat hepatocytes was about one-eighteenth of the rate of incorporation into urea. Thus the purine nucleotide cycle cannot provide most of the ammonia needed in urea synthesis for the carbamoyl phosphate synthase reaction (EC 2.7.2.5). On the other hand, contrary to the view expressed by McGivan & Chappell [(1975) FEBS Lett. 52, 1--7], the experiments support the view that hepatic glutamate dehydrogenase can supply the required ammonia.     

Isolation of pure myocardial subcellular organelles.

A procedure is outlined for the isolation of three pure myocardial subcellular fractions by sucrose gradient ultracentrifugation. The purity of the sarcolemmal (SL) and sarcoplasmic reticulum (SR) fragments and mitochondria were documented by marker enzyme assays and SL purity by electronmicroscopy.     

Immunological and biochemical analysis of a null mutant of α-glycerolphosphate dehydrogenase from Drosophila melanogaster

Summary A Drosophila null mutant(BO-1-4) of -glycerolphosphate dehydrogenase induced by ethylmethane sulfonate(EMS) was analyzed by double immunodiffusion, enzyme immuno-inactivation, immunoelectrophoresis and two-dimensional electrophoresis. Based on all the immunological evidence, this mutant appears to express no protein that can cross-react with the antiserum specific to -glycerolphosphate dehydrogenase. A protein spot corresponding to -glycerolphosphate dehydrogenase was identified on two-dimensional gels of the soluble fly homogenates. The absence of this protein spot on two-dimensional gels of this null mutant further supported the immunological data. The activities of seven other enzymes in the related metabolic pathways were determined for the mutant and the control Drosophila. The null mutant does not show significant alterations in activities of these enzymes. The relationship between the deficiency of this enzyme and the inability for the sustained flight of the null mutant was discussed in terms of cellular metabolic regulations.Abbreviations used -GPD -glycerolphosphate dehydrogenase (EC 1.1.1.8) - EMS ethylmethane sulfonate - TEMED N,N,N,N-tetramethylene diamine - pI isolectric point - CRM immunological cross-reacting material     

Microbiological quality of frozen cauliflower, corn, and peas obtained at retail markets.

The microbiological quality of blanched frozen cauliflower, cut corn, and peas at the retail level was determined. At 35 degrees C, mean aerobic plate count (APC) values for cauliflower, corn, and peas, respectively, were 30,000, 6,100, and 4,700 per g; at 30 degrees C, the mean APC values were 45,000, 8,500, and 6,800 per g, respectively. Geometric means for coliform, Escherichia coli, and Staphylococcus aureus counts for all three vegetables were less than 10 per g.     

Microbiological quality of some spices and herbs in retail markets.

The microbiological quality of 10 spices or herbs was determined by a national survey at the retail level. Aerobic plate count values for the 10 products ranged from less than 100 to 3.1 X 10(8) per g; mean values of the individual spices or herbs ranged from 1,400 to 820,000 per g. Coliform counts ranged from less than 3 to 1.1 X 10(6) per g; however, mean values were less than 20 per g for all products. Escherichia coli counts ranged from less than 3 to 2,300 per g. Except for celery seed, which had a mean value of 7 per g, all mean values were less than 3 per g. Yeast and mold counts were made for 5 of the 10 products. Mean values were generally low; the highest mean (290 per g) was obtained for cinnamon.     

The cDNA for rat selenoprotein P contains 10 TGA codons in the open reading frame

Selenoprotein P is a plasma protein recently purified and characterized as containing 7.5 +/- 1.0 selenium atoms/molecule as selenocysteine. In rats maintained on a defined diet containing nutritionally adequate amounts of selenate as the sole selenium source, over half the selenium in plasma is accounted for by selenoprotein P. Its cDNA has been cloned from a rat liver library and sequenced. The sequence is highly unusual, containing 10 TGA codons in its open reading frame prior to the TAA termination codon. TGA designates selenocysteine in other selenoproteins, and limited peptide sequencing that included the amino acids encoded by two of the TGA codons verified that they correspond to selenocysteine. The deduced 366-amino acid sequence is histidine- and cysteine-rich and contains 9 of its selenocysteines in the terminal 122 amino acids. Comparison of the deduced amino acid sequence of selenoprotein P with those of other selenoprotein reveals no significant similarities. Selenoprotein P represents a new class of selenoproteins and is the first protein described with more than 1 selenocysteine in a single polypeptide chain. The primary structure of selenoprotein P suggests that it might be responsible for some of the antioxidant properties of selenium.     

Point mutations in the DNA polymerase gene of human cytomegalovirus that result in resistance to antiviral agents.

Three independently isolated mutants of human cytomegalovirus strain AD169 were found to be resistant to ganciclovir at a 50% effective dose of 200 microM. Phosphorylation of ganciclovir was reduced 10-fold in mutant-infected cells compared with AD169-infected cells. All three mutants were also determined to be resistant to the nucleotide analogs (S)-1-[(3-hydroxy-2- phosphonylmethoxy)propyl]adenine (HPMPA) and (S)-1-[(3-hydroxy-2-phosphonylmethoxy)propyl]cytosine (HPMPC) and hypersensitive to thymine-1-D-arabinofuranoside (AraT). Single base changes resulting in amino acid substitutions were demonstrated in the nucleotide sequence of the DNA polymerase gene of each mutant. The polymerase mutation contained in one of the mutants was transferred to the wild-type AD169 background. Ganciclovir phosphorylation in cells infected with the recombinant virus produced by this transfer was found to be equivalent to that of AD169-infected cells. The ganciclovir resistance of the recombinant was reduced fourfold compared with that of the parental mutant; however, the recombinant remained resistant to HPMPA and HPMPC and hypersensitive to AraT. The ganciclovir resistance of the mutants therefore appears to result from mutations in two genes: (i) a kinase which phosphorylates ganciclovir and (ii) the viral DNA polymerase.     

Low concentrations of hydrogen sulphide alter monoamine levels in the developing rat central nervous system.

The central nervous system is one of the primary target organs for hydrogen sulphide (H2S) toxicity; however, there are limited data on the neurotoxic effects of low-dose chronic exposure on the developing nervous system. Levels of serotonin and norepinephrine in the developing rat cerebellum and frontal cortex were determined following chronic exposure to 20 and 75 ppm H2S during perinatal development. Both monoamines were altered in rats exposed to 75 ppm H2S compared with controls; serotonin levels were significantly increased at days 14 and 21 postnatal in both brain regions, and norepinephrine levels were significantly increased at days 7, 14, and 21 postnatal in cerebellum and at day 21 in the frontal cortex. Exposure to 20 ppm H2S significantly increased the levels of serotonin in the frontal cortex at day 21, whereas levels of norepinephrine were significantly reduced in the frontal cortex at days 14 and 21, and at day 14 in the cerebellum.     

Achlorhydria induced changes in gastrin, somatostatin, H+/K(+)-ATPase and carbonic anhydrase in the sheep.

Gastrin, somatostatin, H+/K(+)-ATPase and carbonic anhydrase are principal elements of acid secretion. We investigated in the conscious sheep the effect of 24 h omeprazole (an H+/K(+)-ATPase inhibitor) infusion on these elements at the level of synthesis, storage and secretion. Omeprazole inhibited acid secretion-pH increased from 3.0 to 7.1 at 24 h. Plasma amidated and glycine extended gastrin increased 3-fold while the ratio of amidated to glycine extended gastrins (4:1) remained unchanged. Despite the increase in circulating gastrin, antral gastrin concentration and mRNA did not change significantly. Gastrin-17 (amidated and glycine extended) was the predominant form in the circulation and antrum, although there were preferential increases in larger forms following omeprazole treatment. Omeprazole had no effect on somatostatin mRNA or peptide levels in the fundus. Similarly, plasma somatostatin remained unchanged. However, antral somatostatin increased significantly (63%) following omeprazole treatment accompanied by a 4-fold increase in its mRNA. Fundic H+/K(+)-ATPase mRNA was unchanged but a significant increase (87%) in carbonic anhydrase II mRNA was observed. Omeprazole induced hypergastrinaemia occurred without a measurable reduction in storage or increased synthesis of gastrin at 24 h. Increased antral somatostatin synthesis and storage may result from stimulation by plasma gastrin on antral D cells, independent of acid. The rise in carbonic anhydrase II mRNA in the absence of any change in H+/K(+)-ATPase mRNA may reflect the differential sensitivity of the genes encoding these two enzymes to the stimulatory action of gastrin.