1,721,021 research outputs found
FUNCTIONAL RESIDUES AT THE ACTIVE SITE OF HORSE LIVER PHOSPHOPANTOTHENOYLCYSTEINE DECARBOXYLASE.
No full textHorse liver phosphopantothenoylcysteine decarboxylase (EC 4.1.1.36) is rapidly inactivated by N-acetoacetylation with diketene following a pseudo-first-order kinetics: the presence of substrate quantitatively protects against this inactivation. Histidine photo-oxidation with methylene blue or rose bengal brings about the total loss of activity. These results indicate the presence of functional lysyl and histidyl groups at the active site of the enzyme. The substrate sulphydryl group is essential for enzyme activity. Enzymatic decarboxylation is proposed to result from a combined action of the keto group of the enzyme-bound pyruvate protonated by an essential histidine and a protonated amino group of a lysine
PERFUSION OF UREMIC BLOOD ULTRAFILTRATE ON UNCOATED CHARCOAL
No full textA granular uncoated charcoal removes from uremic blood ultrafiltrates many chemical species that are not removed by dialysis. Charcoal treatment dramatically improves the general condition of the patients and normalizes their blood pressure. To obtain a rapid depuration, the initial treatment should be intensive (at least 16 daily treatments) and the effects prolonged over time by once-a-week charcoal treatment between two standard hemodialyses. Biogel P2 chromatography documents well all the events of a depurative treatment that cannot be monitored by hematochemical analyses
Effect of pH salt and temperature on the association of recombinant wt amidase from Sulfolobus solfataricus and on its mutant Y41C.
No full textMolecular and biochemical characterization of the recombinant amidase from hyperthermophilic archaeon Sulfolobus solfataricus
No full textWe have cloned, sequenced, and overexpressed in Escherichia coli the amidase gene from the hyperthermophilic archaeon Sulfolobus solfataricus (strain MT4). The recombinant thermophilic protein was expressed as a fusion protein with an N-terminus six-histidine-residue affinity tag. The enzyme, the first characterized archaeal amidase, is a monomer of 55,784 daltons, enantioselective, and active on 2- to 6-carbon aliphatic amides and on many aromatic amides, over the pH range 4-9 and at temperatures from 60 degrees to 95 degreesC. The S. solfataricus amidase belongs to the class of amidases that share a characteristic signature, GGSS(S/G)GS, located in the central region of the protein, and which show remarkable variability in their individual substrate specificities, can hydrolyze aliphatic or aromatic substrates, and share a large invariance of their primary structure
Cadmium-induced apoptosis and necrosis in human osteoblasts: Role of caspases and mitogen-activated protein kinases pathways
No full textCadmium is a widespread environmental pollutant which induces severe toxic alterations, including osteomalacia and osteoporosis, likely by estrogen receptor-dependent mechanisms. Indeed, cadmium has been described to act as an endocrine disruptor and its toxicity is exerted both in vivo and in vitro through induction of apoptosis and/or necrosis by not fully clarified intracellular mechanism(s) of action. Aim of the present study was to further investigate the molecular mechanism by which cadmium might alter homeostasis of estrogen target cells, such as osteoblast homeostasis, inducing cell apoptosis and/or necrosis. Human osteoblastic cells (hFOB 1.19) in culture were used as an in vitro model to characterize the intracellular mechanisms induced by this heavy metal. Cells were incubated in the presence/absence of 10-50 mu M cadmium chloride at different times and DNA fragmentation and activation of procaspases-8 and -3 were induced upon CdCl2 treatment triggering apoptotic and necrotic pathways. Addition of caspase-8 and -3 inhibitors (Z-IETD-FMK and Z-DQMD-FMK) partially blocked these effects. No activation of procaspase-9 was observed. To determine the role of mitogen-activated protein kinases (MAPK) in these events, we investigated c-jun N-terminal kinase (JNK), p38 and extracellular signal-regulated protein kinase (ERK1/2) phosphorylation which were activated by 10 mu M CdCl2. Chemical inhibitors of JNK, p38, and ERK1/2, SP600125, SB202190, and PD98059, significantly reduced the phosphorylation of the kinases and blunted apoptosis. In contrast, caspase inhibitors did not reduce the cadmium-induced MAPK phosphorylation, suggesting an independent activation of these pathways. In conclusion, at least 2 pathways appear activated by cadmium in osteoblasts: a direct induction of caspase-8 followed by activation of caspase-3 and an indirect induction by phosphorylation of ERK1/2, p38, and JNK MAPK triggering activation of caspase-8 and -3. (J. Endocrinol. Invest. 35: 198-208, 2012) (C) 2012, Editrice Kurti
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