Activation versus Induction of Foetal Enzymes during Culture: a Problem Illustrated by Increased Uridine Diphosphate Glucuronic Acid-5-Hydroxytryptamine Glucuronosyltransferase Activity through Organ Culture on Rafts and Paper Strips

1976 ◽  
Vol 4 (6) ◽  
pp. 1071-1072 ◽  
Author(s):  
JULIAN E. A. LEAKEY ◽  
AGNES M. DONALD
Keyword(s):  
Organ Culture ◽  
Glucuronic Acid ◽  
Uridine Diphosphate

scholarly journals Some properties of the uridine diphosphate glucuronyltransferase activity synthesizing thio-β-d-glucuronides

1973 ◽  
Vol 131 (1) ◽  
pp. 139-147 ◽  
Author(s):  
H. P. A. Illing ◽  
G. J. Dutton
Keyword(s):  
Enzyme Activity ◽  
Phenolic Compounds ◽  
Organ Culture ◽  
Tissue Distribution ◽  
Glucuronic Acid ◽  
Uridine Diphosphate ◽  
Intracellular Location ◽  
Gunn Rats ◽  
Optimum Ph

1. Some properties of the UDP-glucuronyltransferase synthesizing thio-β-d-glucuronides were investigated and compared with those of the enzyme synthesizing the O-glucuronides of analogous phenols. 2. Enzyme activity was generally similar for both classes of substrate in tissue distribution, intracellular location, optimum pH, perinatal development and induction by organ culture or by phenobarbital. 3. Certain differences were noted between the two types of activity in behaviour on storage and on activation, in kinetic behaviour and in distribution between Wistar and Gunn rats; the Gunn rats were not deficient in hepatic UDP-glucuronyltransferase activity towards o-aminothiophenol. 4. These differences are no greater than those exhibited in the synthesis of various O-glucuronides; therefore thiolic substrates could compete in vivo with phenolic compounds for access to the UDP-glucuronyltransferase complex as well as for UDP-glucuronic acid.

scholarly journals Enzymic transfer of glucose and xylose from uridine diphosphate glucose and uridine diphosphate xylose to bilirubin by untreated and digitonin-activated preparations from rat liver

1972 ◽  
Vol 129 (3) ◽  
pp. 619-633 ◽  
Author(s):  
J. Fevery ◽  
P. Leroy ◽  
K. P. M. Heirwegh
Keyword(s):  
Enzyme Activities ◽  
Metal Ion ◽  
Glucuronic Acid ◽  
Uridine Diphosphate ◽  
No Formation ◽  
Cell Extracts ◽  
Straight Line ◽  
Standard Assay ◽  
Diphosphate Glucose ◽  
Liver Homogenates

1. Digitonin-treated and untreated homogenates, cell extracts and washed microsomal preparations from liver of Wistar R rats are capable of transferring sugar from UDP-glucose or UDP-xylose to bilirubin. No formation of bilirubin glycosides occurred with UDP-galactose or d-glucose, d-xylose or d-glucuronic acid as the sources of sugar. 2. Procedures to assay digitonin-activated and unactivated bilirubin UDP-glucosyltransferase and bilirubin UDP-xylosyltransferase were developed. 3. In digitonin-activated microsomal preparations the transferring enzymes had the following properties. Both enzyme activities were increased 2.5-fold by pretreatment with digitonin. They were optimum at pH6.6–7.2. Michaelis–Menten kinetics were followed with respect to UDP-glucose. In contrast, double-reciprocal plots of enzyme activity against the concentration of UDP-xylose showed two intersecting straight-line sections corresponding to concentration ranges where either bilirubin monoxyloside was formed (at low UDP-xylose concentrations) or where mixtures of both the mono- and di-xyloside were synthesized (at high UDP-xylose concentrations). Both enzyme activities were stimulated by Mg2+; Ca2+ was slightly less, and Mn2+ slightly more, stimulatory than Mg2+. Of the activities found in standard assay systems containing Mg2+, 58–78% (substrate UDP-glucose) and 0–38% (substrate UDP-xylose) were independent of added bivalent metal ion. Double-reciprocal plots of the Mg2+-dependent activities against the concentration of added Mg2+ were linear. 4. In comparative experiments the relative activities of liver homogenates obtained with UDP-glucuronic acid, UDP-glucose and UDP-xylose were 1:1.5:2.7 for untreated preparations and 1:0.29:0.44 after activation with digitonin. 5. Bilirubin UDP-glucuronyltransferase was protected against denaturation by human serum albumin, whereas bilirubin UDP-xylosyltransferase was not. 6. Digitonin-treated and untreated liver homogenates from Gunn rats were inactive in transferring sugar to bilirubin from UDP-glucuronic acid (in agreement with the work of others), UDP-glucose or UDP-xylose.

Uridine diphosphate β-glucuronic acid. A new substrate for β-glucuronidase

1970 ◽  
Vol 201 (2) ◽  
pp. 375-377 ◽  
Author(s):  
Indrajit Das ◽  
Mark A. Wentworth ◽  
Hiroyuki Ide ◽  
Hsien Gieh Sie ◽  
William H. Fishman
Keyword(s):  
Glucuronic Acid ◽  
Uridine Diphosphate

A DFT study on the catalytic mechanism of UDP-glucose dehydrogenase

2010 ◽  
Vol 88 (8) ◽  
pp. 804-814 ◽  
Author(s):  
WenJuan Huang ◽  
Jorge Llano ◽  
James W. Gauld
Keyword(s):  
Pathogenic Bacteria ◽  
Catalytic Mechanism ◽  
Glucuronic Acid ◽  
Glucose Dehydrogenase ◽  
Polarizable Continuum Model ◽  
Uridine Diphosphate ◽  
Reaction Field ◽  
Enzyme Active Site ◽  
Site Model ◽  
Diphosphate Glucose

Uridine 5′-diphosphate glucuronic acid (UDPGlcUA) is a key intermediary metabolite in many species, including pathogenic bacteria and humans. It is biosynthesized from UDP-glucose (UDPGlc) by uridine diphosphate glucose dehydrogenase (UDPGlcDH) via a twofold two-electron–one-proton oxidation that successively transforms the 6-hydroxymethyl of glucopyranose into a formyl, and the latter into the final carboxylic function. The catalytic mechanism of UDPGlcDH was investigated using a large enzyme active-site model in combination with the B3LYP method and the polarizable continuum model (IEF-PCM) self-consistent reaction field. The latter was used to correct for the long-range electrostatic effect of the protein environment. The overall mechanism consists of four catalytic steps: (i) NAD+-dependent oxidation of glucose to glucuronaldehyde, (ii) nucleophilic addition of Cys260–SH to glucuronaldehyde to form a 6-thiohemiacetal intermediate, (iii) NAD+-dependent oxidation of the 6-thiohemiacetal to form a 6-thioester intermediate, and finally, (iv) hydrolysis of the 6-thioester to give glucuronic acid. In addition, this study also provides insight into the debated roles of Lys204 and Asp264, and the most likely protonation state of a reactive Michaelis complex of UDPGlcDH.

Increased Glucuronidation of Bilirubin in Man and Rat by Administration of Antipyrine (Phenazone)

1974 ◽  
Vol 46 (4) ◽  
pp. 511-518
Author(s):  
M. L'E. Orme ◽  
L. Davies ◽  
A. Breckenridge
Keyword(s):  
Protein Content ◽  
Glucuronic Acid ◽  
Serum Bilirubin ◽  
Microsomal Protein ◽  
Affinity Constant ◽  
Maximal Velocity ◽  
Uridine Diphosphate ◽  
Bilirubin Concentration ◽  
Contrast Administration ◽  
Significant Difference

1. Antipyrine in a dose of 3·2 mmol (600 mg) daily for 6 weeks produced a significant fall in both total and unconjugated serum bilirubin concentrations in six patients with Gilbert's syndrome. The maximum reduction in serum bilirubin concentration was seen after 2 weeks of treatment. 2. In the rat, administration of antipyrine in doses of 0·42 and 1·27 mmol 24 h−1 kg−1 (80 and 240 mg 24 h−1 kg−1) for 84 h caused a significant increase in the apparent maximal velocity (Vmax.) for the glucuronidation of bilirubin by liver microsomal preparations when the concentration of either uridine diphosphate glucuronic acid (UDPGA) or bilirubin was altered. There was no significant difference between the apparent Vmax. values attained with the two doses of antipyrine in either set of experiments. Neither the microsomal protein content nor the apparent affinity constant (Km) was altered in these studies. 3. In contrast, administration of phenobarbitone in doses of 0·34 mmol 24 h−1 kg−1 (80 mg 24 h−1 kg−1) caused a significant increase in the microsomal protein content but there was no significant change in the values for the apparent Vmax. or apparent Km for the glucuronidation of bilirubin with various concentrations of both UDPGA and bilirubin.

THE ACID-SOLUBLE NUCLEOTIDES OF SALMON LIVER

1958 ◽  
Vol 36 (5) ◽  
pp. 465-473 ◽  
Author(s):  
H. Tsuyuki ◽  
Violet M. Chang ◽  
D. R. Idler
Keyword(s):  
Low Temperature ◽  
Anion Exchange ◽  
Rat Liver ◽  
Glucuronic Acid ◽  
Anion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Uridine Diphosphate ◽  
Carbohydrate Storage ◽  
Diphosphate Glucose

The acid-soluble nucleotides of spring salmon liver have been separated by anion-exchange chromatography at low temperature and characterized. Under these conditions the relatively labile uridine-5′-diphosphate nucleotides of acetylglucosamine, galactose, and glucuronic acid were obtained intact, a fact that is further substantiated by the complete absence of uridine-5′-diphosphate. The occurrence of these uridine diphosphate compounds and the absence of uridine diphosphate glucose is discussed in relation to the previously postulated role of inositol as a carbohydrate storage product. A new peptide-containing nucleotide, succinoadenosine-5′-phosphosulphate (peptide), was found in the fraction which immediately follows adenosine-5′-diphosphate. The parent base of this nucleotide, succinoadenine, was also isolated. The nucleotide pattern is simpler than that reported by other investigators for rat liver and wheat.

scholarly journals The effect of steroids and nucleotides on solubilized bilirubin uridine diphosphate glucuronyltransferase

1970 ◽  
Vol 119 (3) ◽  
pp. 437-445 ◽  
Author(s):  
B. P. F. Adlard ◽  
G. H. Lathe
Keyword(s):  
Enzyme Activity ◽  
Glucuronic Acid ◽  
Liver Microsomes ◽  
Fold Increase ◽  
Rat Liver Microsomes ◽  
Uridine Diphosphate ◽  
Competitive Effects ◽  
Solubilized Enzyme ◽  
High Concentrations ◽  
Solubilized Form

1. It was confirmed that bilirubin glucuronyltransferase can be obtained in solubilized form from rat liver microsomes. 2. Michaelis–Menten kinetics were not followed by the enzyme with bilirubin as substrate when the bilirubin/albumin ratio was varied. High concentrations of bilirubin were inhibitory. 3. The Km for UDP-glucuronic acid at the optimum bilirubin concentration was 0.46mm. 4. Low concentrations of Ca2+ were inhibitory in the absence of Mg2+ but stimulatory in its presence; the converse applied for EDTA. 5. UDP-N-acetylglucosamine and UDP-glucose enhanced conjugation by untreated, but not by solubilized microsomes. 6. The apparent 9.5-fold increase in activity after solubilization was probably due to the absence of UDP-glucuronic acid pyrophosphatase activity in the solubilized preparation. 7. The activation of solubilized enzyme activity by ATP was considered to be a result of chelation of inhibitory metal ions. 8. The solubilized enzyme activity was inhibited by UMP and UDP. The effect of UMP was not competitive with respect to UDP-glucuronic acid. 9. A number of steroids inhibited the solubilized enzyme activity. The competitive effects of stilboestrol, oestrone sulphate and 3β-hydroxyandrost-5-en-17-one, with respect to UDP-glucuronic acid, may be explained on an allosteric basis.

scholarly journals The binding of oxidized and reduced nicotinamide–adenine dinucleotides to bovine liver uridine diphosphate glucose dehydrogenase

1974 ◽  
Vol 141 (3) ◽  
pp. 667-673 ◽  
Author(s):  
Paul A. Gainey ◽  
Charles F. Phelps
Keyword(s):  
Glucuronic Acid ◽  
Glucose Dehydrogenase ◽  
Gel Filtration ◽  
Bovine Liver ◽  
Uridine Diphosphate ◽  
Protein Fluorescence ◽  
Diphosphate Glucose ◽  
Ph Dependent ◽  
Uridine Diphosphate Glucose Dehydrogenase ◽  
Biphasic Profile

The binding of NAD+and NADH to bovine liver UDP-glucose dehydrogenase was studied by using gel-filtration and fluorescence-titration methods. The enzyme bound 0.5mol of NAD+and 2 mol of NADH/mol of subunit at saturating concentrations of both substrate and product. The dissociation constant for NADH was 4.3μm. The binding of NAD+to the enzyme resulted in a small quench of protein fluorescence whereas the binding of NADH resulted in a much larger (60–70%) quench of protein fluorescence. The binding of NADH to the enzyme was pH-dependent. At pH8.1 a biphasic profile was obtained on titrating the enzyme with NADH, whereas at pH8.8 the titration profile was hyperbolic. UDP-xylose, and to a lesser extent UDP-glucuronic acid, lowered the apparent affinity of the enzyme for NADH.

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