1,721,103 research outputs found
Bromopyruvate for the affinity labeling of a residue near the carboxylate binding site of 6-phosphogluconate dehydrogenase
No full text
Degradation of aromatic compounds by Trichosporon Sp
No full textWhen cultured in liquid media, samples of the yeast Trichosporon grow readily and degrade phenol; glutamate was found to stimulate both fungal growth and phenol catabolism, with a distinctive lag. In addition, this same strain grows in the presence of 2-chloro-phenol and 2-methyl, 4-chlorophenol (which are also degraded) and in the presence of 2-nitrophenol, 4-nitrophenol, 2,4-dinitrophenol, which are either degraded or not, as in the case of 4-nitrophenol. The kinetics of both growth and of aromatic catabolism is suggestive of inductive phenomena for key metabolic enzymes
Glyoxylate for affinity labelling of 6-phosphogluconate dehydrogenase
No full textIn order to find a new reagent for the affinity labelling, 6-phosphogluconate dehydrogenase was treated with glyoxylate, a versatile metabolite with a carboxyl and a reactive aldehydic group. High concentrations of glyoxylate inhibit the enzyme, while in the presence of the reducing agent cyanoborohydride, the enzyme is irreversibly inactivated by only millimolar glyoxylate. This indicates the formation of a Schiff base between the aldehydic group of glyoxylate and one enzyme lysine residue. The kinetics and substrate competition suggest that inactivation is due to affinity labelling. In the first step the inhibitor carboxylic group binds to the substrate carboxyl binding site, and in the second slower step the aldehydic group binds a nearby lysine. We have also found that other enzymes are inactivated by the combined actions of glyoxylate and cyanoborohydride, with a saturation kinetics. Hence, glyoxylate can be helpful to identify specific lysines at the carboxyl binding sites in pro..
Inhibitors behaviour agree with the 3-dimensional structure of the active site in 6-phosphogluconate dehydrogenases
No full text6-Phosphogluconate dehydrogenase and its crystal structures
No full text6-Phosphogluconate dehydrogenase (6PGDH; EC 1.1.1.44) catalyses the
oxidative decarboxylation of 6-phosphogluconate to ribulose 5-phosphate in
the context of the oxidative part of the pentose phosphate pathway. Depending
on the species, it can be a homodimer or a homotetramer. Oligomerization plays
a functional role not only because the active site is at the interface between
subunits but also due to the interlocking tail-modulating activity, similar to that
of isocitrate dehydrogenase and malic enzyme, which catalyse a similar type of
reaction. Since the pioneering crystal structure of sheep liver 6PGDH, which
allowed motifs common to the -hydroxyacid dehydrogenase superfamily to be
recognized, several other 6PGDH crystal structures have been solved, including
those of ternary complexes. These showed that more than one conformation
exists, as had been suggested for many years from enzyme studies in solution.
It is inferred that an asymmetrical conformation with a rearrangement of one of
the two subunits underlies the homotropic cooperativity. There has been
particular interest in the presence or absence of sulfate during crystallization.
This might be related to the fact that this ion, which is a competitive inhibitor
that binds in the active site, can induce the same 6PGDH configuration as in the
complexes with physiological ligands. Mutagenesis, inhibitors, kinetic and
binding studies, post-translational modifications and research on the enzyme
in cancer cells have been complementary to the crystallographic studies.
Computational modelling and new structural studies will probably help to refine
the understanding of the functioning of this enzyme, which represents a
promising therapeutic target in immunity, cancer and infective diseases. 6PGDH
also has applied-science potential as a biosensor or a biobattery. To this end, the
enzyme has been efficiently immobilized on specific polymers and nanoparticles.
This review spans the 6PGDH literature and all of the 6PGDH crystal structure
data files held by the Protein Data Bank
Acanthosis nigricans-insulin resistance Type A syndrome: analysis of restriction fragments length polymorphisms at the insulin receptor locus
No full textWe have identified two sisters (12 and 17 years old) affected by acanthosis
nigricans-insulin resistance (AN-IR) type A syndrome and
Type 2 (non-insulin-dependent) diabetes mellitus. They presented
with acanthosis nigricans, marked hyperinsulinaemia and severe insulin
resistance, Type 2 diabetes, no antibodies to the insulin receptor,
obesity and virilisation without other endocrine diseases. Both
parents and paternal grandmother had Type 2 diabetes. In AN-IR
type A syndrome a primary defect of insulin receptor is supposed.
The availability of cloned DNA (cDNA) for the human insulin receptor
allows examination of the possible role of this gene in this
syndrome. Therefore we analysed restriction fragments length polymorphisms
(RFLP) for the insulin receptor gene in different members
of this family, including diabetic and non diabetic subjects.
DNA extracted from white blood cells was digested by seven restriction
enzymes, analysed by Southern blotting technique, using a
eDNA probe for the insulin receptor of 4.2 kilobases. Insulin receptor
DNA fragments appeared the same in all the examined subjects.
No association of any RFLP was noted with the syndrome. Therefore,
in this family, RFLPs for the insulin receptor gene were uninformative
in evaluating the role of this gene in AN-IR type A syndrome,
nevertheless the obtained results exclude its marked alterations
in the investigated patients
Active site labeling of erythrocyte transglutaminase by o-phthalaldehyde
No full textTissue-type transglutaminase is inactivated in a time-dependent way during incubation with submillimolar concentrations of o-phthalaldehyde, with affinity labeling kinetics. The rate of inactivation by the reagent is greatly enhanced in the presence of the essential enzyme cofactor calcium and is decreased by GTP, an allosteric inhibitor. A fluorescent isoindole derivative is formed during the modification apparently through crosslinkage of active site Cys 277 to a lysine residue. These data and the quenching of fluorescence by addition of calcium ions suggest that the enzyme active site is directly involved in the inactivation process
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