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Biomedical subjects

L Uotila

Publications and source records attributed to L Uotila.

At least 37 records · Page 2Linked to original sources
PubMed1983

Inhibition of vitamin K-dependent carboxylase in vitro by cefamandole and its structural analogs.

Clinical use of the beta-lactam cephalosporins has resulted in a number of reports of vitamin K-responsive bleeding abnormalities. The effects of these antibiotics on the activities of vitamin K-dependent enzymes in vitro were therefore assessed. The activity of vitamin K-dependent carboxylase was inhibited by 50% by 6-10 mM cefamandole or structurally analogous cephalosporins. The inhibition was noncompetitive with respect to the pentapeptide substrate, vitamin KH2, and CO2. Vitamin K epoxide reductase was less sensitive to and only partially inhibited by cefamandole and the analogs. The possible physiologic significance of the inhibition of these enzymes in clinical hemorrhagic problems was considered.

Animals↗
PubMed1982

Characterization of vitamin K-dependent carboxylase from the livers from the adult ox and dicoumarol-treated calf.

The properties of the microsomal vitamin K-dependent carboxylase from the livers of the adult ox and dicoumarol-treated calf were investigated. The enzymes from both sources utilized glutamic residues of synthetic peptides as substrates and could be solubilized with Triton X-100 similarly to the enzyme from vitamin K-deficient rat liver. Under the optimal assay conditions, the microsomes from calf liver had peptide carboxylase activity comparable with that of the rat liver microsomes and 6.5-fold that of adult ox liver microsomes. The apparent Km for reduced vitamin K and the ionic strength optima of the calf and adult ox enzyme clearly differ from those of the rat enzyme. Pyridoxal phosphate activated the adult ox carboxylase only slightly, whereas the calf enzyme was activated by pyridoxal phosphate as effectively as was the enzyme from the vitamin K-deficient rat. Mn2+ activated the adult ox enzyme 9-fold and calf enzyme 22-fold under optimal conditions (no KCl). Three other divalent metal cations (Ca2+, Ba2+, and Mg2+) activated the adult ox and calf enzymes to about half the extent caused by Mn2+, KCl inhibited this activation. The vitamin K-dependent carboxylase from the dicoumarol-treated calf is apparently more tightly bound to the microsomal membrane than is the adult ox enzyme. In many other respects (pH optimum), temperature optimum, Km values for peptide substrate, substrate specificity, inhibitor effects), the properties of the adult ox and calf enzymes resemble closely those of the rat enzyme.

Age Factors↗
PubMed1982

Recent findings in understanding the biological function of vitamin K.

Vitamin K is required for the posttranslational modification of liver precursors of prothrombin, blood coagulation Factors VII, IX, and X and additional proteins of undetermined functions in plasma and other tissues. This modification involves the formation of gamma-carboxyglutamic acid, an acidic amino acid needed for the interaction of these proteins with calcium ions. The vitamin is a cofactor of a unique microsomal carboxylase which requires the reduced form of vitamin K, CO2, and molecular oxygen. The vitamin is apparently oxidized to its 2,3-epoxide during the carboxylation reaction. Liver microsomes also contain an enzyme which catalyzes the reduction of the epoxide to the vitamin, vitamin K epoxide reductase, and a number of vitamin K reductases. The epoxide reductase appears to be the site of the anticoagulant action of 4-hydroxycoumarins, commonly used as oral anticoagulants.

1-Carboxyglutamic Acid↗
PubMed1981

The relation of the soluble thiamine triphosphatase activity of various rat tissues to nonspecific phosphatases.

Polyacrylamide gel electrophoresis was used to investigate the relation of the soluble thiamine triphosphatase activity of various rat tissues to other phosphatases. This technique separated the thiamine triphosphatase of rat brain, heart, kidney, liver, lung, muscle and spleen from alkaline phosphatase (EC 3.1.3.1), acid phosphatase (EC 3.1.3.2) and other nonspecific phosphatase activities. In contrast, the hydrolytic activity for thiamine triphosphate in rat intestine moved identically with alkaline phosphatase in gel electrophoresis. Thiamine triphosphatase from rat liver and brain was also separated from alkaline phosphatase and acid phosphatase by gel chromatography on Sephadex G-100. This gave an apparent molecular weight of about 30,000 and a Stokes radius of 2.5 nanometers for brain and liver thiamine triphosphatase. The intestinal thiamine triphosphatase activity of the rat was eluted from the Sephadex G-100 column as two separate peaks (with apparent molecular weights of over 200,000 and 123,000) which exactly corresponded to the peaks of alkaline phosphatase. The isoelectric point (pI) of the brain thiamine triphosphatase was 4.6 (4 degrees C). The partially purified thiamine triphosphatase from brain and liver was highly specific for thiamine triphosphate. The results suggest that, apart from the intestine, the rat tissues studied contain a specific enzyme, thiamine triphosphatase (EC 3.6.1.28). The specific enzyme is responsible for most of the thiamine triphosphatase activity in these tissues. Rat intestine contains a high thiamine triphosphatase activity but all of it appears to be due to alkaline phosphatase.

Acid Phosphatase↗
PubMed1980

Product inhibition studies of human liver formaldehyde dehydrogenase.

The steady-state kinetic mechanism of human liver formaldehyde dehydrogenase (formaldehyde:NAD+ oxidoreductase (glutathione-formylating), EC 1.2.1.1) was investigated by product inhibition of the forward and the reverse reactions catalyzed by the enzyme. The results are compatible with a mechanism which contains the random addition to the enzyme of NAD+ and S-hydroxymethylglutathione (the adduct of glutathione and formaldehyde), or NADH and S-formylglutathione, and free glutathione as the allosteric activator of the enzyme (Uotila, L. and Mannervik, B. (1979) Biochem. J. 177, 869-878).

Aldehyde Oxidoreductases↗
PubMed1980

Transient effect of L-tryptophan in progressive myoclonus epilepsy without Lafora bodies: clinical and electrophysiological study.

A double-blind crossover trial with 2 g L-tryptophan and placebo was carried out with five familial and two sporadic patients with progressive myoclonus epilepsy (PME) without Lafora bodies. L-Tryptophan improved the clinical condition in six out of seven patients. Clinical improvement in ambulation, myoclonic jerks, and general condition was most evident. The change was statistically significant. In visual assessment of EEGs, the amount of paroxysmal activity and dysrhythmia of the background activity decreased in six out of seven patients on L-tryptophan. The quantitative EEG revealed a decrease in the power bands of theta, alpha, and beta activity in five of six patients on the second day of L-tryptophan treatment. In familial PME cases, the responses were consistently beneficial. With long-term L-tryptophan therapy, the effect disappeared or was even reversed in three of seven patients after 3 to 4 weeks. These findings indicate that therapy with serotonin precursors is worthy of further trial in PME and that deficient tryptophan metabolism may play a part in the etiology of PME without Lafora bodies.

Adolescent↗
PubMed1979

Glutathione thiol esterases of human red blood cells. Fractionation by gel electrophoresis and isoelectric focusing.

The number and the substrate specificities of glutathione thiol esterases of human red blood cells have been investigated by gel electrophoresis and isoelectric focusing and staining methods devised for the location of these enzymes on gels. Several glutathione thiol esterase forms, both unspecific (with respect to the S-acyl group of the substrate) and specific were found. Electrophoresis on both polyacrylamide and agarose gels resolved three enzyme components with apparently similar substrate specificity. Isoelectric focusing in liquid column separated two unspecific thiol esterase components with S-lactoylglutathione (pI = 8.4) and S-propionylglutathione (pI = 8.1) as the best substrates, respectively, and two specific enzymes, S-formylglutathione hydrolase (pI = 5.2) and S-succinylglutathione hydrolase (pI = 9.0). Isoelectric focusing on polyacrylamide gel resolved nine unspecific glutathione thiol esterase bands (between pH values 7.0 and 8.4). Partially purified glyoxalase II (S-2-hydroxyacylglutathione hydrolase, EC 3.1.2.6) from erythrocytes or liver still gave three components on electrophoresis and several activity bands on gel electrofocusing. These results indicate that human red cells contain at least four separate glutathione thiol esterases. Glyoxalase II, one of these enzymes, apparently occurs in multiple forms. These were neither influenced by preptreatment of the samples with neuraminidase or thiols nor were interconvertible during the fractionations.

Electrophoresis, Polyacrylamide Gel↗
PubMed1979

A steady-state-kinetic model for formaldehyde dehydrogenase from human liver. A mechanism involving NAD+ and the hemimercaptal adduct of glutathione and formaldehyde as substrates and free glutathione as an allosteric activator of the enzyme.

The steady-state kinetics of formaldehyde dehydrogenase from human liver have been explored. Non-linearities were obtained in v-versus-v[S] plots. It was necessary and sufficient to consider two reactants of the equilibrium mixture of formaldehyde, glutathione and their hemimercaptal adduct for a complete description of the kinetics. A random sequential reaction scheme is proposed in which adduct and beta-NAD+ are the substrates. In addition, glutathione can bind to an allosteric regulatory site and only the glutathione-containing enzyme is considered productive. Various alternative reaction models were examined but no simple alterative was superior to the model chosen. The discrimination was largely based on results of non-linear regression analysis. Several S-substituted glutathione derivatives were tested as activators or inhibitors of the enzyme, but all were without effect. Thio-NAD+, nicotinamide--hypoxanthine dinucleotide and 3-acetylpyridine-adenine dinucleotide could substitute for beta-NAD+ as the nucleotide substrate. alpha-NAD+ and ADP-ribose were competitive inhibitors with respect to beta-NAD+ and non-competitive with glutathione and the adduct. When used simultaneously, the inhibitors were linear competitive versus each other, indicating a single nucleotide-binding site or, if more than one, non-co-operative binding sites.

Aldehyde Oxidoreductases↗
PubMed1979

Separation of the isoenzymes of glyoxalase I from human red blood cells by electrophoresis and isoelectric focusing on polyacrylamide gel and by ion exchange chromatography.

Methods have been devised for the separation of the isoenzymes of glyoxalase I(S-lactoylglutathione methylglyoxal-lyase (isomerizing), EC 4.4.1.5) from human red blood cells by electrophoresis and electrofocusing on polyacrylamide gel slabs. Three different staining methods were used for the location of the enzyme. Three electrophoretic phenotypes of the enzyme were resolved, the fast and slow types with one band and the intermediate type with three glyoxalase I activity bands. In gel electrofocusing (pH gradient 3.5-9.5) two glyoxalase I activity bands were found for all electrophoretic types. In electrofocusing on gel with a narrow pH gradient, at least four separate enzyme components were resolved for the fast and slow electrophoretic types and at least six components for the intermediate type. The phenotypes could be distinguished correspondingly to the electrophoretic results. Preparative separation of the isoenzymes were achieved by ion exchange chromatography on DEAE-Sephacel but gel chromatography on Sephadex G-100 gave the same elution volume for all enzyme phenotypes. This corresponds to an apparent molecular weight of about 47 000.

Chromatography, Ion Exchange↗