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The present study investigates the biotransformation of glucose with uridine monophosphate (UMP) to obtain sugar nucleotide, UDP-glucose (UDPG), by the dried cells of Candida saitoana KCTC7249. The biotransformation was optimized by varying the concentrations of substrates and phosphate ion. UDPG (24 mM) was biotransformed from 200 mM glucose and 37.5 mM UMP by dried cells of C. saitoana. The glucose yields about 64% UDP-glucose, based on UMP concentration. The addition of glucose-1-phosphate to the reaction mixture accelerated the formation of UDPG from a concentration of UMP. The structure of UDP-glucose obtained was determined with 13C NMR and FAB mass spectra. These results indicate that the yeast-dried cells could be used for the production of nucleotide sugars for donor molecules of complex carbohydrate synthesis.
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The deficiency of uridine-5'-monophosphate synthase is inherited as an autosomal recessive trait in Holstein-Friesian cattle. Heterozygotes are characterized by half normal activity of uridine monophosphate synthase, as measured in erythrocytes. Matings between heterozygotes for the deficiency have produced 31 pregnancies lasting at least one month, but only 23 full-term calves, including 9 homozygous normal and 14 heterozygotes. That the eight embryos were not carried to term is consistent with the probability of a homozygous-deficient genotype and are lost around 40 d of gestation. Although the homozygous deficiency of uridine monophosphate synthase is lethal embryonically in cattle, homozygotes for the analogous human condition have been born alive.
A partial deficiency of uridine monophosphate synthase has been discovered among Holstein dairy cattle wherein affected animals have half the normal activity of this enzyme and are heterozygous for a condition presumed to be lethal in the homozygous recessive state. A random sample of 287 dairy bulls, active in artificial insemination organizations, were assayed for their activity of uridine monophosphate synthase. Four bulls displayed reduced enzyme activity (heterozygotes) corresponding to a 1.4% incidence and were closely related to a common ancestor, Bull-X. Testing a total of 75 male descendants of Bull-X has uncovered 22 heterozygotes. Heterozygotes include 5 sons, 15 grandsons, 1 a double grandson, and 2 great grandsons of Bull-X. Reproductive failure from this enzyme deficiency has been minimal because matings between close relatives have been infrequent. If the incidence of heterozygotes active in artificial insemination increases, avoidance of matings between close relatives will become an inadequate safety measure to maintain reproductive fitness. Random mating of male heterozygotes to a female population with 2% incidence of heterozygotes may imply an additional .5% embryonic loss relative to matings involving bulls that are normal for uridine monophosphate synthase.
Interaction between genotype at locus for uridine monophosphate synthase and parity was reexamined for 305-d mature equivalent milk and fat production and calving interval. Three lactations for cows of known genotype defined two data files corresponding to different sampling procedures. The first data file selected by herd contained records on 23 cows that were deficient in activity of uridine monophosphate synthase and on 1081 cows with normal activity; the second data file contained records on 19 cows with deficient activity and on 17 cows with normal activity, all daughters of bulls known to be heterozygous at the locus for uridine monophosphate synthase. Results from the mutually exclusive data files were determined independently and then pooled. Genotype and parity interaction, tested against mean square error, was not supported for any of the three traits. Mean differences of the two genotypes, averaged across data files, were, for first, second, and third parities, respectively, 360, 568, and 299 kg for milk; 6, 9, and 8 kg for fat; and 22, 9, and 12 d for calving interval. Main effect of genotype indicated a 417-kg advantage for heterozygotes but 17-d longer calving interval. The main effect of genotype on milk production adjusted for calving interval was 343 kg but was nonsignificant.
An inherited deficiency of uridine-5'-monophosphate synthase has recently been discovered in dairy cattle. The possibility was explored that this deficiency might be detected by low amounts of erythrocyte 3-N-ribosyluric acid, since it is synthesized by uridine monophosphate synthase. Ribosyluric acid was lower in six dairy cows with the deficiency compared to six controls, paired on the basis of age, stage of lactation, and diet (.66 vs. 1.38 mumol/ml). However, ribosyluric acid concentration measured in 50 normal cows showed a wide range (.28 to 2.35 mumol/ml), with nine lower than the mean for the deficient animals. This extensive variability may limit the usefulness of ribosyluric acid in diagnosing the inherited deficiency of uridine monophosphate synthase.
Cows with a partial deficiency of uridine monophosphate synthase have elevated orotate in their milk and urine. Variability of orotate was assessed in two such cows monitored biweekly for two complete lactations. Concentrations of milk and urinary orotate showed extensive variation during lactation, but coefficients of variation were similar for deficient and normal cows. Orotate averaged 500 micrograms/ml milk for deficient cows (versus a normal of 80 micrograms/ml), and a threshold of 200 micrograms/ml distinguished normal from deficient cows. Deficient cows had low milk orotate upon initiation of lactation and exhibited a latency of 1 to 7 wk to attain that threshold. The deficiency also resulted in a lactation-induced orotic aciduria. Orotate averaged 32.2 micrograms/ml urine for deficient cows (versus a normal of 7.3 micrograms/ml), and a threshold of 15 micrograms/ml urine differentiated the animals. Latency was 3 to 18 wk for deficient cows to exceed that threshold. Total orotate output and orotate concentration were elevated in milk and urine of lactating cows deficient for uridine monophosphate synthase. The output of orotate was predominantly in milk rather than urine for both deficient and normal cows. Additionally, orotate was elevated in blood of deficient cows when they were lactating.
Deficiency of uridine monophosphate synthase (DUMPS) is a monogenic autosomal recessive disorder in cattle, resulting in early embryonic death of homozygous offspring. To identify the mutation responsible for DUMPS, liver RNA from identified, DUMPS heterozygous animals from the Holstein and Red Holstein breeds was reverse transcribed. Amplification of cDNA with sequence-specific primers and subsequent sequencing of the PCR products revealed a mutation (C-->T) with the loss of an AvaI site at codon 405, resulting in a premature stop codon with a truncated C-terminal catalytic subunit of the protein. A direct DNA test based on PCR was developed and subsequently tested on 102 animals. Complete concurrence of deficiency of UMPS and the presence of the described point mutation in heterozygous animals was observed, thus confirming this point mutation as the basic defect in DUMPS cattle.
The SOC8 gene was isolated as an extragenic suppressor of cdc8 mutant cells. It has been suggested that SOC8 is allelic with the URA6 gene which was originally identified as a uridine monophosphate kinase. In this article, we describe the purification of the uridine monophosphate kinase from a yeast Saccharomyces cerevisae strain that overproduces the activity 8-fold. The protein was purified through Fast-Flow Q-Separose, phosphocellulose, blue-agarose, and fast protein liquid chromatography Superose 12 columns, and appears homogeneous by sodium dodecyl sulfate-polyacrylamide gel analysis. The uridine monophosphate kinase contains a single polypeptide with a molecular weight of 25,000, as evidence by both sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration analysis. The amino acid composition has also been determined. Substrate specificity studies show that the relative activity of nucleoside monophosphates is in order of UMP greater than dUMP, and to a lesser extent, dTMP, GMP, and dGMP. The Km and Vm of UMP, dUMP, and dTMP have been determined.
Uridine monophosphate synthase plays an important role in pyrimidine synthesis, converting orotic acid to uridine 5' monophosphate. In cattle, UMPS deficiency is inherited as a monogenic autosomal recessive trait. While heterozygous carriers are phenotypically normal, homozygotes are lethal in utero (Shanks et al. 1992). UMPS has been mapped to human chromosome 3q13 (Qumsiyeh et al. 1989), sheep chromosome 1q (Burkin et al. 1993) and cattle chromosome 1q31 (Ryan et al. 1994). In the present study we used a cattle genomic probe to localize UMPS to river buffalo chromosomes by fluorescence in situ hybridization.
Red cell uridine monophosphate kinase polymorphism has been studied on a total of 915 individuals from two different areas of Italy (Milan and Rome). The two groups of about the same size show very similar gene frequencies. The UMPK2 allele in the pooled sample has a frequency of 2.8% which is significantly lower than those observed in the two other Caucasian populations so far examined.
Milk orotate concentration was tested for 1451 registered and grade Holsteins from 23 herds. Cows with milk orotate concentration exceeding 200 micrograms/ml were blood-tested for uridine monophosphate synthase activity. Thirty-four cows were diagnosed as deficient in this enzyme, which is necessary for the metabolic conversion of orotate to uridine 5'-monophosphate. To control for genetics, a subset of 33 deficient cows and their 145 normal paternal half-sisters were also analyzed. Milk yield at 305 d, adjusted for age and milking frequency, was significantly affected by the stage of lactation at which the test was done, by the parity of the cow, and by the interaction of the parity of the cow with its diagnosis of uridine monophosphate synthase in both analyses. Twenty-five older, deficient cows yielded the most milk (8118); 8 first-calf, deficient cows the least milk (6810); and 1386 normal cows intermediate amounts. Deficiency effect and interaction of parity with deficiency were not significant for fat. Interaction of parity with deficiency was significant for calving interval. Older, deficient cows had the longest calving intervals; first calf, deficient cows had the shortest calving intervals; and normal cows had intermediate calving intervals. However, deficiency was not associated with milk yield in analysis that considered calving interval as a covariate for milk.
This study aimed to demonstrate the effect of uridine monophosphate (UMP) on the consolidation of a brightness-discrimination reaction after topical application of this RNA precursor into the hippocampus, the neocortex or the mesencephalic reticular formation (MRF). Thirty minutes before the rats started their training in a Y-chamber, UMP was injected into each animal through cannula implanted into the particular brain area. When injected into hippocampus or MRF UMP exerted no influence on acquisition, but after epidural UMP injection an impairment of acquisition was observed. After intrahippocampal or epidural UMP application the retention test carried out 48 hour after training showed a significant improvement in retention performance, whereas UMP injection into MRF showed no influence on retention. Consequently, the retention-improving effect of UMP is probably not induced by activation of ascending neuronal systems.
Cattle heterozygous for deficiency of uridine monophosphate synthase required more breeding services per calving when mated to other heterozygotes than did matings of normal cattle. Gestation length, number of breeding services per calving, and days from breeding to pregnancy examination were monitored on 759 complete gestations, 76 false-positive pregnancy diagnoses, 14 false-negative pregnancy diagnoses, and 413 negative pregnancy diagnoses in a dairy herd between 1983 and 1987. For complete gestations, 15 heterozygote x heterozygote matings required 3.13 +/- 0.37 breeding services per calving, which was significantly more than the 2.05 +/- 0.05 breeding services required for normal x normal matings; gestation length and days from breeding to pregnancy examination were similar between mating types. For false-positive pregnancy diagnoses, females diagnosed pregnant, but subsequently found not to be pregnant, 5 heterozygote x heterozygote matings averaged 51 +/- 23 days of gestation, which was less than the 93 +/- 6 days required for 71 normal matings; services and days from breeding to pregnancy examination were similar between mating types. All false-negative pregnancy diagnoses, females diagnosed not pregnant but later found to be pregnant, were made on cattle with normal matings. For negative pregnancy diagnoses, heterozygous matings averaged 0.3 more breeding services per examination than normal matings.