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W L Yang

Publications and source records attributed to W L Yang.

18 recordsLinked to original sources
PubMed1995

Differential biochemical regulation of the URA7- and URA8-encoded CTP synthetases from Saccharomyces cerevisiae.

The URA7- and URA8-encoded CTP synthetases (EC 6.3.4.2, UTP:ammonia ligase (ADP-forming) are functionally overlapping enzymes responsible for the biosynthesis of CTP in the yeast Saccharomyces cerevisiae. URA8-encoded CTP synthetase was purified to apparent homogeneity by ammonium sulfate fractionation of the cytosolic fraction followed by chromatography with Q-Sepharose, Affi-Gel Blue, Mono Q, and Superose 6. The subunit molecular mass (67 kDa) of purified URA8-encoded CTP synthetase was in good agreement with the predicted size of the URA8 gene product. Antibodies raised against a fusion protein constructed from the coding sequences of the URA8 gene and expressed in Escherichia coli reacted with purified URA8-encoded CTP synthetase. Native URA8-encoded CTP synthetase existed as a dimer which oligomerized to a tetramer in the presence of its substrates UTP and ATP. Maximum URA8-encoded CTP synthetase activity was dependent on Mg2+ ions (Ka = 2.4 mM) and 2-mercaptoethanol at the pH optimum of 7.5. The enzyme followed saturation kinetics toward UTP (Km = 74 microM), ATP (Km = 22 microM), and glutamine (Km = 0.14 mM). GTP stimulated (Ka = 26 microM) URA8-encoded CTP synthetase activity 12-fold. CTP potently inhibited (IC50 = 85 microM) URA8-encoded CTP synthetase activity and, in addition, caused the dependence of activity toward UTP to become cooperative. The URA8-encoded CTP synthetase and the previously purified URA7-encoded CTP synthetase differed significantly with respect to several biochemical properties including turnover number, pH optimum, substrate dependences, and sensitivity to inhibition by CTP. The URA7-encoded CTP synthetase mRNA was 2-fold more abundant when compared with URA8-encoded CTP synthetase mRNA. Both CTP synthetase isoforms were maximally expressed in the exponential phase of growth.

Base Sequence↗
PubMed1995

Phosphorylation of CTP synthetase from Saccharomyces cerevisiae by protein kinase C.

Phosphorylation of CTP synthetase (EC 6.3.4.2, UTP:ammonia ligase (ADP-forming)) from Saccharomyces cerevisiae protein kinase C was examined. Using pure CTP by synthetase as a substrate, protein kinase C activity was dose- and time-dependent and required calcium, diacylglycerol, and phosphatidylserine for full activation. Protein kinase C activity was also dependent on the concentration of CTP synthetase. Protein kinase C phosphorylated CTP synthetase on serine and threonine residues in vitro whereas the enzyme was primarily phosphorylated on serine residues in vivo. Phosphopeptide mapping analysis of CTP synthetase phosphorylated in vitro and in vivo indicated that the enzyme was phosphorylated on more than one site. Most of the phosphopeptides derived from CTP synthetase phosphorylated in vivo were the same as those derived from CTP synthetase phosphorylated by protein kinase C in vitro. The stoichiometry of the phosphorylation of native CTP synthetase was 0.4 mol of phosphate/mol of enzyme whereas the stoichiometry of the phosphorylation of alkaline phosphatase-treated CTP synthetase was 2.2 mol of phosphate/mol of enzyme. This indicated that CTP synthetase was purified in a phosphorylated state. Phosphorylation of CTP synthetase resulted in a 3-fold activation in enzyme activity whereas alkaline phosphatase treatment of CTP synthetase resulted in a 5-fold decrease in enzyme activity. Overall, the results reported here were consistent with the conclusion that CTP synthetase was regulated by protein kinase C phosphorylation.

Adenosine Triphosphate↗
PubMed1994

Purification and characterization of CTP synthetase, the product of the URA7 gene in Saccharomyces cerevisiae.

In the yeast Saccharomyces cerevisiae, CTP synthetase [EC 6.3.4.2; UTP:ammonia ligase (ADP-forming)] is the product of the URA7 gene. CTP synthetase was purified 503-fold to apparent homogeneity from cells bearing the URA7 gene on a multicopy plasmid that directed a 10-fold overproduction of the enzyme. The purification procedure included ammonium sulfate fractionation of the cytosolic fraction followed by chromatography with Sephacryl 300 HR, Q-Sepharose, Affi-Gel Blue, and Superose 6. The N-terminal amino acid sequence of purified CTP synthetase was identified and aligned perfectly with the deduced sequence of the URA7 gene. The minimum subunit molecular mass (68 kDa) of purified CTP synthetase was in good agreement with the size (64.7 kDa) of the URA7 gene product. Antibodies were raised against a maltose-binding protein-CTP synthetase fusion protein which immunoprecipitated CTP synthetase from wild-type cells. Immunoblot analysis was used to identify CTP synthetase in wild-type cells and cells bearing the URA7 gene on a multicopy plasmid. The results of gel filtration chromatography indicated that the size of native CTP synthetase was consistent with a dimeric structure for the enzyme. CTP synthetase oligomerized to a tetramer in the presence of its substrates UTP and ATP. Maximum CTP synthetase activity was dependent on magnesium ions (4 mM) and 2-mercaptoethanol at the pH optimum of 8.0. CTP synthetase exhibited positive cooperative kinetics with respect to UTP and ATP and negative cooperative kinetics with respect to glutamine and GTP. CTP synthetase was potently inhibited by the product CTP which also increased the positive cooperativity of the enzyme toward UTP.(ABSTRACT TRUNCATED AT 250 WORDS)

Base Sequence↗
PubMed1994

Use of synthetic lethal mutants to clone and characterize a novel CTP synthetase gene in Saccharomyces cerevisiae.

In the pyrimidine biosynthetic pathway, CTP synthetase catalyses the conversion of uridine 5'-triphosphate (UTP) to cytidine 5'-triphosphate (CTP). In the yeast Saccharomyces cerevisiae, the URA7 gene encoding this enzyme was previously shown to be nonessential for cell viability. The present paper describes the selection of synthetic lethal mutants in the CTP biosynthetic pathway that led us to clone a second gene, named URA8, which also encodes a CTP synthetase. Comparison of the predicted amino acid sequences of the products of URA7 and URA8 shows 78% identity. Deletion of the URA8 gene is viable in a haploid strain but simultaneous presence of null alleles both URA7 and URA8 is lethal. Based on the codon bias values for the two genes and the intracellular concentrations of CTP in strains deleted for one of the two genes, relative to the wild-type level, URA7 appears to be the major gene for CTP biosynthesis. Nevertheless, URA8 alone also allows yeast growth, at least under standard laboratory conditions.

Amino Acid Sequence↗
PubMed1992

Structural basis of the in vivo induction of micronuclei.

The structural basis of the in vivo induction of micronuclei was examined with CASE, a structure-activity relational method. The CASE program identified a number of structures associated with this activity. When used to predict the activity of chemicals not included in the learning set, these structural determinants gave a concordance in excess of 83%. The existence of a structural basis for the induction of micronuclei will permit an investigation of the mechanistic basis of this phenomenon.

Databases, Factual↗
PubMed1991

[Surgical treatment of superior mesenteric artery syndrome].

Fourty-two patients with superior mesenteric artery syndrome were treated surgically. Among the patients, 24 were male and 18 female (mean age, 38 years). The courses of the disease varied from 1 to 10 years. Operations included amputation of the ligament of Treitz (14 patients), gastrojejunostomy (2), subtotal gastrectomy and gastro jejunostomy (1), duodenojejunostomy (6), anterior repositioning of the duodenum (7), and circular drainage operation of the duodenum (12). Two patients underwent anterior repositioning of the duodenum and gastrojejunostomy. In patients without relief of symptoms such as vomiting after operation, circular drainage of the duodenum was performed. All the 42 patients were successfully treated. The authors discussed 6 kinds of operations, their indications, and the advantages and disadvantages. Emphasis was given to the anterior repositioning of the duodenum and its circular drainage.

Adolescent↗
PubMed1989

[Protective effect of extracts from Aloe vera L. var. chinensis (Haw.) Berg. on experimental hepatic lesions and a primary clinical study on the injection of in patients with hepatitis].

The injection(10-15 ml/kg/d, ip x 4), total glycoside (125-225 mg/kg/d, ip x (3-4); 600 mg/kg/d, ig x 3) and crystal III (120 mg/kg/d, ip x 4) of Aloe vera var. chinensis were found to be effective in lowering the elevated sGPT induced by CCl4, thioacetamide and D-aminogalactose in mice or rats. It was also observed that these agents could protect hepatic cells from the CCl4-induced injury. When dogs were given in with Aloe injection of 0.1 ml/kg/d x 180, no toxicity was noted. The total effective sGPT-lowering rate of Aloe injection on 38 patients of chronic hepatitis with positive HBsAg was 86.8%.

Alanine Transaminase↗
PubMed1989

[Histologic distribution of anti-glioma monoclonal antibody SZ-39 in human xenograft brain tumors].

131I-labeled anti-glioma monoclonal antibody SZ-39 was injected intraperitoneally into nude mice bearing xenograft human glioma, ependymoma and intracranial metastatic adenocarcinoma. Subsequent gamma-imaging and assessment of radioactivity showed selective accumulation of antibody in the glioma. Quantitative autoradiography supported the results of radiolocalization observed in vivo in different brain tumors and normal tissues. It was also shown that antibody SZ-39 was specially localized to viable glioma cells and closely associated with their cell membrane. Yet, it was not bound to fibrous or necrotic areas. 131I-normal mouse IgG was distributed in a nonspecific pattern in the glioma. These studies demonstrated the specificity of SZ-39 in vivo which may be useful in clinical diagnostic guiding and treatment.

Adenocarcinoma↗
PubMed1987

Presence of hepatitis B virus DNA in serum of surface-antigen-seronegative immunocompromised patients.

We compared the prevalence of serum hepatitis B virus (HBV) DNA in cancer patients who were immunocompromised with that in healthy subjects. Testing positive for serum HBV DNA were 27 of the 232 (11.6%) surface-antigen-negative cancer patients and 7 of the 382 (1.8%) surface-antigen-negative healthy subjects. These data suggest that the negative serology, at least in immunocompromised individuals, is probably not sufficient to exclude HBV infection.

DNA, Viral↗
PubMed1987

Hepatitis type B virus DNA in patients receiving hemodialysis: correlation with other HBV serological markers.

Possible presence of hepatitis type B virus (HBV) was assessed in 239 end-stage renal failure patients who were receiving long-term maintenance hemodialysis (average 30.8 months; duration: 1-94 months), and who had not shown any other symptom of HBV infection. Their HBV serological markers, including HBV DNA, were evaluated together with those of normal control individuals. HBV surface antigen (HBsAg) was detected in 42 of the 239 dialysis patients, 15 of whom also positive for HBV DNA (mean +/- SD = 56.2 +/- 23.7 pg/100 microliters of serum). HBV DNA was also found in 22 of the 197 (11.2%) dialysis patients who were negative for HBsAg, with an average of 36.2 +/- 19.0 pg/100 microliters of serum. This rate of detecting HBV DNA in HBV seronegative dialysis individuals was significantly higher than the rate of 1.83% found among healthy HBsAg(-) individuals. Among these 22 dialysis patients who were HBsAg(-) but HBV DNA(+), 15 were found to possess antibodies against HBsAg (anti-HBs) and/or antibody against HBV e antigen (anti-HBe). These data suggested that the absence of serum HBV antigen or the presence of antibodies against HBV markers might not be sufficient to identify possible HBV infection in immunocompromised hosts such as hemodialysis patients.

Adult↗