Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.27.5 (RNase)
 17,967 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Hepatic ethanol metabolism generates the reactive intermediate, acetaldehyde, which binds to proteins. The binding of acetaldehyde to purified enzymes was determined in order to ascertain whether such binding altered their catalytic functions. [14C]Acetaldehyde was incubated with alcohol dehydrogenase, glucose-6-phosphate dehydrogenase, lactate dehydrogenase and RNase A, each at 37 degrees C (pH 7.4). In some reactions, sodium cyanoborohydride was included for stabilization of Schiff bases, formed as a result of the reaction between acetaldehyde and the amino groups of the enzymes. Portions of each reaction mixture were removed for determination of stable and total (stable plus borohydride-reducible) adducts. Alcohol dehydrogenase and lactate dehydrogenase were not inhibited by adduct formation. Glucose-6-phosphate dehydrogenase and RNase, the activities of which depend on a lysine residue at their catalytic sites, were inhibited in a dose- and time-dependent manner. The degree of inhibition directly correlated with total adduct formation. Phosphate, known to inhibit binding to the active site lysine of RNase, prevented the inhibition of catalytic activity caused by adduct formation. These findings indicate that the binding of acetaldehyde to lysine at the catalytic site can inhibit enzyme activity.
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PMID:Covalent binding of acetaldehyde selectively inhibits the catalytic activity of lysine-dependent enzymes. 293 8

Ribonuclease A was reacted with [1-13C,1,2-14C]acetaldehyde and sodium cyanoborohydride in the presence or absence of 0.2 M phosphate. After several hours of incubation at 4 degrees C (pH 7.4) stable acetaldehyde-RNase adducts were formed, and the extent of their formation was similar regardless of the presence of phosphate. Although the total amount of covalent binding was comparable in the absence or presence of phosphate, this active site ligand prevented the inhibition of enzymatic activity seen in its absence. This protective action of phosphate diminished with progressive ethylation of RNase, indicating that the reversible association of phosphate with the active site lysyl residue was overcome by the irreversible process of reductive ethylation. Modified RNase was analysed using 13C proton decoupled NMR spectroscopy. Peaks arising from the covalent binding of enriched acetaldehyde to free amino groups in the absence of phosphate were as follows: NH2-terminal alpha amino group, 47.3 ppm; bulk ethylation at epsilon amino groups of nonessential lysyl residues, 43.0 ppm; and the epsilon amino group of lysine-41 at the active site, 47.4 ppm. In the spectrum of RNase ethylated in the presence of phosphate, the peak at 47.4 ppm was absent. When RNase was selectively premethylated in the presence of phosphate, to block all but the active site lysyl residues and then ethylated in its absence, the signal at 43.0 ppm was greatly diminished, and that arising from the active site lysyl residue at 47.4 ppm was enhanced. These results indicate that phosphate specifically protected the active site lysine from reaction with acetaldehyde, and that modification of this lysine by acetaldehyde adduct formation resulted in inhibition of catalytic activity.
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PMID:The binding of acetaldehyde to the active site of ribonuclease: alterations in catalytic activity and effects of phosphate. 365 Nov 78

The effects of acute and chronic treatments with ethanol and acute treatments with an ethanol metabolite, acetaldehyde, on proopiomelanocortin (POMC) mRNA expression were compared with those of these agents on the secretion of a POMC gene product, beta-endorphin (beta-EP) peptide. The level of POMC mRNA in cultured cells was determined using an RNase protection assay, and the accumulation of immunoreactive beta-EP (IR-beta-EP) peptide in the culture medium was measured by radioimmunoassay. Treatment of hypothalamic cells with 25-, 50-, and 100-mM doses of ethanol or 12.5 and 25 microM acetaldehyde for 3 h increased POMC mRNA levels. The stimulatory effect of ethanol on POMC mRNA levels lasted for a period of 12 h, although the percentage increase of the ethanol-stimulated mRNA level was gradually reduced over time. Acute treatments with ethanol and acetaldehyde also elevated IR-beta-EP secretion from the cultured neurons for a period of 12 h, and the IR-beta-EP secretory response developed desensitization after 24 h of ethanol incubation. The close association between the ethanol-induced IR-beta-EP secretion and ethanol-regulated POMC mRNA expression suggests that ethanol regulates both secretion and production of beta-EP peptide in the hypothalamic neurons.
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PMID:Comparison of the effects of alcohol and acetaldehyde on proopiomelanocortin mRNA levels and beta-endorphin secretion from hypothalamic neurons in primary cultures. 770 32

Cytochrome P-450 2E1 (CYP2E1) is a readily inducible hemoprotein that catalyzes the oxidation of endogenous compounds and many low molecular weight xenobiotics. As the major component of the microsomal ethanol oxidizing system, it contributes significantly to ethanol metabolism and the formation of the highly reactive metabolite acetaldehyde. The leaky property of this enzyme results in the generation of reactive oxygen species that can induce oxidative stress and cytotoxic conditions deleterious to development. To further investigate the proposed role of CYP2E1 in the etiology of alcohol teratogenesis, the current study focused on the quantification of CYP2E1 in prenatal human brain, a tissue that is highly vulnerable to the damaging effects of ethanol throughout gestation. In microsomal samples prepared from pools of brain tissues, immunoreactive protein was detected by Western blot analysis using enhanced chemiluminescence, whereas functional protein was estimated with an enzymatic assay using p-nitrophenol and an electrochemical detection system. CYP2E1 transcript was consistently detected in RNA samples prepared from individual brain tissues using the ribonuclease protection assay. Quantitative data were collected by scanning densitometry and phosphorimaging technology. There was a dramatic increase in human brain CYP2E1 content around gestational day 50 and a fairly constant level was maintained throughout the early fetal period, until at least day 113. The relatively low levels of the P-450 isoform present in conceptal brain may be sufficient to generate reactive intermediates that elicit neuroembryotoxicity following maternal alcohol consumption.
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PMID:Catalytic activity and quantitation of cytochrome P-450 2E1 in prenatal human brain. 1033 64

Alcohol misusers frequently have difficulties in gait, and various muscle symptoms such as cramps, local pain and reduced muscle mass. These symptoms are common in alcoholic patients and have previously been ascribed as neuropathological in origin. However, biochemical lesions and/or the presence of a defined myopathy occur in alcoholics as a direct consequence of alcohol misuse. The myopathy occurs independently of peripheral neuropathy, malnutrition and overt liver disease. Chronic alcoholic myopathy is characterized by selective atrophy of Type II fibres and the entire muscle mass may be reduced by up to 30%. This myopathy is arguably the most prevalent skeletal muscle disorder in the Western Hemisphere and occurs in approximately 50% of alcohol misusers. Alcohol and acetaldehyde are potent inhibitors of muscle protein synthesis, and both contractile and non-contractile proteins are affected by acute and chronic alcohol dosage. Muscle RNA is also reduced by mechanisms involving increased RNase activities. In general, muscle protease activities are either reduced or unaltered, although markers of muscle membrane damage are increased which may be related to injury by reactive oxygen species. This supposition is supported by the observation that in the UK, alpha-tocopherol status is poor in myopathic alcoholics. Reduced alpha-tocopherol may pre-dispose the muscle to metabolic injury. However, experimental alpha-tocopherol supplementation is ineffective in preventing ethanol-induced lesions in muscle as defined by reduced rates of protein synthesis and in Spanish alcoholics with myopathy, there is no evidence of impaired alpha-tocopherol status. In conclusion, by a complex series of mechanisms, alcohol adversely affects skeletal muscle. In addition to the mechanical changes to muscle, there are important metabolic consequences, by virtue of the fact that skeletal muscle is 40% of body mass and an important contributor to whole-body protein turnover.
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PMID:Alcoholic skeletal muscle myopathy: definitions, features, contribution of neuropathy, impact and diagnosis. 1178 53