Biosensors for phenol derivatives using biochemical signal amplification

Two different approaches, both exploiting two enzymes cooperative functioning, to enhance the sensitivity of tyrosinase (PPO) based biosensor for amperometric detection of phenols have been compared. For this purpose, one monoenzyme electrode (PPO) and two bienzyme electrodes (PPO and d-glucose dehydrogenase, GDH; PPO and horseradish peroxidase, HRP) were constructed using agar-agar gel as enzyme immobilization matrix. The biosensors responses for l-tyrosine detection were recorded at −50 mV versus saturated calomel electrode (SCE). The highest sensitivity (74 mA M⁻¹) was observed for the PPO-GDH couple, while that recorded for PPO-HRP couple system was only 32 times higher than that measured for monoenzyme electrode (0.01 mA M⁻¹). The ability of the PPO-, PPO-GDH-, PPO-HRP-based biosensors to assay phenols was demonstrated by quantitative determination of phenol, 1,2-dihydroxybenzene, 1,3-dihydroxybenzene, 1,4-dihydroxybenzene, 2-amino-3 (4-hydroxyphenyl) propanoic acid, 2-hydroxytoluene, 3-hydroxytoluene, 4-hydroxytoluene, 4-clorophenol, 3-clorophenol, 2-clorophenol, 4-hydroxybenzoic acid.     

Tracking X‐ray‐derived redox changes in crystals of a methylamine dehydrogenase/amicyanin complex using single‐crystal UV/Vis microspectrophotometry

X‐ray exposure during crystallographic data collection can result in unintended redox changes in proteins containing functionally important redox centers. In order to directly monitor X‐ray‐derived redox changes in trapped oxidative half‐reaction intermediates of Paracoccus denitrificans methylamine dehydrogenase, a commercially available single‐crystal UV/Vis microspectrophotometer was installed on‐line at the BioCARS beamline 14‐BM‐C at the Advanced Photon Source, Argonne, USA. Monitoring the redox state of the intermediates during X‐ray exposure permitted the creation of a general multi‐crystal data collection strategy to generate true structures of each redox intermediate.     

Synthesis and Characterization of Some New Quinoxalin-2(1H)one and 2-Methyl-3H-quinazolin-4-one Derivatives Targeting the Onset and Progression of CRC with SAR,Molecular Docking,and ADMET Analyses

The pathogenesis of colorectal cancer is a multifactorial process. Dysbiosis and the overexpression of COX-2 and LDHA are important effectors in the initiation and development of the disease through chromosomal instability, PGE2 biosynthesis, and induction of the Warburg effect, respectively. Herein, we report the in vitro testing of some new quinoxalinone and quinazolinone Schiff’s bases as: antibacterial, COX-2 and LDHA inhibitors, and anticolorectal agents on HCT-116 and LoVo cells. Moreover, molecular docking and SAR analyses were performed to identify the structural features contributing to the biological activities. Among the synthesized molecules, the most active cytotoxic agent, (6d) was also a COX-2 inhibitor. In silico ADMET studies predicted that (6d) would have high Caco-2 permeability, and %HIA (99.58%), with low BBB permeability, zero hepatotoxicity, and zero risk of sudden cardiac arrest, or mutagenicity. Further, (6d) is not a potential P-gp substrate, instead, it is a possible P-gpI and II inhibitor, therefore, it can prevent or reverse the multidrug resistance of the anticancer drugs. Collectively, (6d) can be considered as a promising lead suitable for further optimization to develop anti-CRC agents or glycoproteins inhibitors.     

A new mechanism of acyl group transfer in the reaction catalyzed by D-glyceraldehyde-3-phosphate dehydrogenase

D-Glyceraldehyde-3-phosphate dehydrogenase (GAPDH, EC 1.2.1.12) catalyzes the oxidative phosphorylation of its substrate in a two-step reaction. Asa result of the first, oxidativestep, the covalent intermediate where in 3-phosphoglyceroyl moiety is bound to Cys149 of the active center is subjected to nucleophilic attack by inorganic phosphate, but remains resistant to hydrolytic decomposition. This ensures tight coupling of oxidation with phosphorylation in glycolysis. In this article, we present the experimental evidence for the conversion of GAPDH into a form capable of performing the reaction in the absence of inorganic phosphate. The structural basis for this conversion is the oxidation of a cysteine residue (probably Cys 153) into a sulfenic acid derivative under mild conditions to affect the integrity of the essential Cys 149. As a result, an intram olecular transfer of 3-phosphoglyceroyl group from the active center Cys 149 to Cys 153 becomes possible with subsequent hydrolysis of the sulfenyl carboxylate intermediate.     

Biocatalytic reduction system for the production of chiral methyl (R)/(S)-4-bromo-3-hydroxybutyrate

An effective method for producing methyl 4-bromo-3-hydroxybutyrate enantiomers was developed using an engineered protein. Escherichia coli transformant cells containing a mutant β-keto ester reductase (KER-L54Q) from Penicillium citrinum and a cofactor-regeneration enzyme such as glucose dehydrogenase (GDH) or Leifsonia sp. alcohol dehydrogenase (LSADH) were used to produce methyl (S)-4-bromo-3-hydroxybutyrate from methyl 4-bromo-3-oxobutyrate. On the other hand, the production of methyl (R)-4-bromo-3-hydroxybutyrate was achieved by asymmetric reduction of methyl 4-bromo-3-oxobutyrate with a mutant phenylacetaldehyde reductase (PAR-HAR1) from Rhodococcus sp. ST-10.     

Carbon Nanotubes‐Ionic Liquid and Chloropromazine Modified Electrode for Determination of NADH and Fabrication of Ethanol Biosensor

Potential cycling was used for oxidation of chloropromazine and producing an electroactive redox couples which strongly adsorbed on the electrode surface modified with carbon nanotubes and ionic liquid nanocomposite. The modified electrode shows excellent electrocatalytic activity toward NADH oxidation. The differential pulse voltammetry detection provided high sensitivity, 0.5835 A M^?1, low detection limit, 80 nM at concentration range up to 20 μM. An ethanol biosensor was also developed by immobilizing alcohol dehydrogenase enzyme onto nanocomposite. Differential pulse voltammetric detection of ethanol gives linear responses over the concentration range 40 μM–1.5 mM with detection limit 5 μM and sensitivity 1.97 μA mM^?1.     

Norcholanic acids as substrates for recombinant 3β-hydroxysteroid dehydrogenase and progesterone 5β-reductase, enzymes of the 5β-cardenolide biosynthesis

The conversion of 23-nor-5,20(22)E-choladienic acid-3β-ol and other intermediates of the putative norcholanic acid pathway of cardenolide biosynthesis by recombinant 3β-hydroxysteroid dehydrogenase from Digitalis lanata in dehydrogenation and reduction reactions was investigated. 23-nor-4,20(22)E-choladienic acid-3-one was found to be a substrate of recombinant progesterone 5β-reductases from D. lanata and Arabidopsis thaliana. The role of various substrates in cardenolide biosynthesis is discussed.     

Assay of Serum Glutamic-Oxaloacetic Transaminase Using Malate Dehydrogenase Preparation Obtained from Streptomyces aureofaciens

Abstract

A modified spectrophotometric method for serum glutamic-oxaloacetic transaminase (SGOT) assay was developed. A crude cell-free extract from Streptomyces aureofaciens which showed a high level of malate dehydrogenase (MDH) activity (E.C. 1.1.1.37) was used as the enzymatic indicator. The lyophilized microbial preparation was used without previous purification and was quite stable under refrigeration for one year. Serum sample assays using both the method utilizing the crude cell extract and an enzymatic commercial kit showed good correlation.     

Effects of He－Ne laser irradiation on enzyme activity in milking Goats＇ seminal plasma

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