The competitive inhibition of yeast alcohol dehydrogenase by 2,2,2-trifluoroethanol

Taber, Richard L.

doi:10.1016/s0307-4412(98)00073-9

Cited by 18 publications

(14 citation statements)

References 21 publications

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“…Each well contained 110 mM ethanol or ethanol- d 6 , 0.125–4.00 mM NAD + , 0.024 unit/mL (0.45 nM) YADH, and 300 g/L crowding agent. The YADH stock solution was prepared by adding 8 μL of a 2 g/L enzyme solution to 5992 μL of 1 g/L BSA, which served to stabilize and maintain enzyme activity . Addition of ethanol initiated the enzymatic reaction, but adding the reagents in a different order did not have an effect on the resulting rate.…”

Section: Methodsmentioning

confidence: 99%

“…The YADH stock solution was prepared by adding 8 μL of a 2 g/L enzyme solution to 5992 μL of 1 g/L BSA, which served to stabilize and maintain enzyme activity. 39 Addition of ethanol initiated the enzymatic reaction, but adding the reagents in a different order did not have an effect on the resulting rate. The assay was repeated using 550 mM isopropanol or isopropanol-d 8 in place of ethanol and 0.25−12.5 mM NAD + .…”

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confidence: 95%

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Effects of Macromolecular Crowding on Alcohol Dehydrogenase Activity Are Substrate-Dependent

2016

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Enzymes operate in a densely packed cellular environment that rarely matches the dilute conditions under which they are studied. To better understand the ramifications of this crowding, the Michaelis-Menten kinetics of yeast alcohol dehydrogenase (YADH) were monitored spectrophotometrically in the presence of high concentrations of dextran. Crowding decreased the maximal rate of the reaction by 40% for assays with ethanol, the primary substrate of YADH. This observation was attributed to slowed release of the reduced β-nicotinamide adenine dinucleotide product, which is rate-limiting. In contrast, when larger alcohols were used as the YADH substrate, the rate-limiting step becomes hydride transfer and crowding instead increased the maximal rate of the reaction by 20-40%. This work reveals the importance of considering enzyme mechanism when evaluating the ways in which crowding can alter kinetics.

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Section: Methodsmentioning

confidence: 99%

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confidence: 95%

Effects of Macromolecular Crowding on Alcohol Dehydrogenase Activity Are Substrate-Dependent

2016

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“…There are several studies on inhibition mechanism of ADH from obtained various sources. For example, some reports showed that 2,2,2-trifluoroethanol, (Taber, 1998) thiol compound (pyrazoles) (Li et al, 2010) and 4,4' dithiodipyridine (Zheng et al, 1997) inhibited the alcohol dehydrogenase. In another study, isolated catechins, and flavonoids from the leaves of Camellia sinensis exhibited yeast alcohol dehydrogenase (ADH) inhibitory activities in the range of IC 50 8.0-70.3μM (Manir et al, 2012).…”

Section: Discussionmentioning

confidence: 99%

Alcohol Dehydrogenase from Sheep Liver: Purification, Characterization and Impacts of Some Antibiotics

Demir¹,

Şengül²,

Erğun³

et al. 2017

jist

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Alcohol dehydrogenase (ADH) is a dimeric enzyme in which each subunit of the enzyme has a Zn 2+ metal-containing catalytic domain and a cofactor-binding domain. This enzyme converts the alcohol to aldehyde. The present article focuses on the purification, characterization and in vitro effects of some antibiotics on alcohol dehydrogenase from sheep liver. ADH was purified with specific activity of 0.5 U/mg proteins and approximately 52.03-fold from sheep liver by DEAE-Sephadex A-50 ion exchange chromatography and gel filtration on Sephadex G-100. The subunit and the natural molecular weights of the enzyme were determined by gel filtration and SDS-PAGE 80.49 and 38.16 kDa, respectively. The optimum ionic strenght, temperature and, pH of ADH was 400 mM, 40°C and, 10.5, respectively. The inhibitory effects of the antibiotics were tested at various concentrations. IC 50 values for kanamycin sulfate, amikacin sulfate, gentamicin, lincomycin, and clindamycin were found to be 43. 31, 36.47, 20.38, 18.73 and 1.31 mM, respectively.

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“…4). As stated in the previous studies, the binding of iodoacetic acid, N-ethylmaleimide, 4,4′-dithiodipyridine, or 4-hydroxymercury benzoic acid to the enzyme active site are irreversible [23][24][25]. Therefore, the presence of these four inhibitors during cultivation of living R. oryzae prevented further enzyme-substrate binding.…”

Section: 2-diazole and 222-trifluoroethanol As The Competitive Admentioning

confidence: 90%

“…Many studies on the in vitro inhibition of yeast ADH (purified/partially purified) have been reported. Use of a substrate analog such as 2,2,2-trifluoroethanol or modification of ADH conformation by addition of sulfhydryl reagents showed significant inhibition of ADH activity [23][24][25]. Nevertheless, little is known about R. oryzae ADH inhibition especially in the in vivo test.…”

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confidence: 99%

In Vivo Regulation of Alcohol Dehydrogenase and Lactate Dehydrogenase in Rhizopus Oryzae to Improve l-Lactic Acid Fermentation

Thitiprasert

Sooksai

Thongchul

2011

Appl Biochem Biotechnol

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Rhizopus oryzae is becoming more important due to its ability to produce an optically pure L: -lactic acid. However, fermentation by Rhizopus usually suffers from low yield because of production of ethanol as a byproduct. Limiting ethanol production in living immobilized R. oryzae by inhibition of alcohol dehydrogenase (ADH) was observed in shake flask fermentation. The effects of ADH inhibitors added into the medium on the regulation of ADH and lactate dehydrogenase (LDH) as well as the production of cell biomass, lactic acid, and ethanol were elucidated. 1,2-diazole and 2,2,2-trifluroethanol were found to be the effective inhibitors used in this study. The highest lactic acid yield of 0.47 g/g glucose was obtained when 0.01 mM 2,2,2-trifluoroethanol was present during the production phase of the pregrown R. oryzae. This represents about 38% increase in yield as compared with that from the simple glucose fermentation. Fungal metabolism was suppressed when iodoacetic acid, N-ethylmaleimide, 4,4'-dithiodipyridine, or 4-hydroxymercury benzoic acid were present. Dramatic increase in ADH and LDH activities but slight change in product yields might be explained by the inhibitors controlling enzyme activities at the pyruvate branch point. This showed that in living R. oryzae, the inhibitors regulated the flux through the related pathways.

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The competitive inhibition of yeast alcohol dehydrogenase by 2,2,2-trifluoroethanol

Cited by 18 publications

References 21 publications

Effects of Macromolecular Crowding on Alcohol Dehydrogenase Activity Are Substrate-Dependent

Effects of Macromolecular Crowding on Alcohol Dehydrogenase Activity Are Substrate-Dependent

Alcohol Dehydrogenase from Sheep Liver: Purification, Characterization and Impacts of Some Antibiotics

In Vivo Regulation of Alcohol Dehydrogenase and Lactate Dehydrogenase in Rhizopus Oryzae to Improve l-Lactic Acid Fermentation

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