Transient kinetic data for partial reactions of alcohol dehydrogenase and simulations of progress curves have led to estimates of rate constants for the following mechanism, at pH 8.0 and 25 degrees C: E in equilibrium E-NAD+ in equilibrium *E-NAD+ in equilibrium E-NAD(+)-RCH2OH in equilibrium E-NAD+-RCH2O- in equilibrium *E-NADH-RCHO in equilibrium E-NADH-RCHO in equilibrium E-NADH in equilibrium E. Previous results show that the E-NAD+ complex isomerizes with a forward rate constant of 620 s-1 [Sekhar, V. C., & Plapp, B. V. (1988) Biochemistry 27, 5082-5088]. The enzyme-NAD(+)-alcohol complex has a pK value of 7.2 and loses a proton rapidly (greater than 1000 s-1). The transient oxidation of ethanol is 2-fold faster in D2O, and proton inventory results suggest that the transition state has a charge of -0.3 on the substrate oxygen. Rate constants for hydride ion transfer in the forward or reverse reactions were similar for short-chain aliphatic substrates (400-600 s-1). A small deuterium isotope effect for transient oxidation of longer chain alcohols is apparently due to the isomerization of the E-NAD+ complex. The transient reduction of aliphatic aldehydes showed no primary deuterium isotope effect; thus, an isomerization of the E-NADH-aldehyde complex is postulated, as isomerization of the E-NADH complex was too fast to be detected. The estimated microscopic rate constants show that the observed transient reactions are controlled by multiple steps.

The binding of NAD+ to liver alcohol dehydrogenase was studied by stopped-flow techniques in the pH range from 6.1 to 10.9 at 25 degrees C. Varying the concentrations of NAD+ and a substrate analogue used to trap the enzyme-NAD+ complex gave saturation kinetics. The same maximum rate constants were obtained with or without the trapping agent and by following the reaction with protein fluorescence or absorbance of a ternary complex. The data fit a mechanism with diffusion-controlled association of enzyme and NAD+, followed by an isomerization with a forward rate constant of 500 s-1 at pH 8: E E-NAD+ *E-NAD+. The isomerization may be related to the conformational change determined by X-ray crystallography of free enzyme and enzyme-coenzyme complexes. Overall bimolecular rate constants for NAD+ binding show a bell-shaped pH dependence with apparent pK values at 6.9 and 9.0. Acetimidylation of epsilon-amino groups shifts the upper pK to a value of 11 or higher, suggesting that Lys-228 is responsible for the pK of 9.0. Formation of the enzyme-imidazole complex abolishes the pK value of 6.9, suggesting that a hydrogen-bonded system extending from the zinc-bound water to His-51 is responsible for this pK value. The rates of isomerization of E-NAD+ and of pyrazole binding were maximal at pH below a pK of about 8, which is attributable to the hydrogen-bonded system. Acetimidylation of lysines or displacement of zinc-water with imidazole had little effect on the rate of isomerization of the E-NAD+ complex.(ABSTRACT TRUNCATED AT 250 WORDS)

A series of 2-o-arylidineaminophenylindoles and their cyclic derivatives (indolo[1,2-c]quinazolines) were synthesized. The reactions occurred under relatively mild conditions and afforded the desired product in good yields. Molecular structures of the synthesized compounds were confirmed by IR, (1)H-NMR,( 13)C-NMR, MS spectra, and elemental analyses. Furthermore, all the final products were screened for in-vitro antibacterial activity against three Gram-positive and three Gram-negative bacteria and also tested for their inhibitory action against three strains of fungi. Compound IIc showed potent activity against all the bacterial (except S. typhimurium) and fungal strains. Especially, compounds IIi and IIj which have isoquinolyl and pyridyl substituents displayed potent antibacterial as well as antifungal activities compared to those of the respective standard drugs Ampicillin and Ketoconazole.

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Background Mycosis on the fall armyworm, Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae), infecting maize was observed in research farm of Regional Agricultural Research Station, Anakapalli from October 2019 to February 2020. Main body High relative humidity (94.87%), low temperature (24.11 °C), and high rainfall (376.1 mm) received during the month of September 2019 predisposed the larval instars for fungal infection and subsequent high relative humidity and low temperatures sustained the infection till February 2020. An entomopathogenic fungus (EPF) was isolated from the infected larval instars as per standard protocol on Sabouraud’s maltose yeast extract agar and characterized based on morphological and molecular analysis. The fungus was identified as Metarhizium (Nomuraea) rileyi based on ITS sequence homology and the strain was designated as AKP-Nr-1. The pathogenicity of M. rileyi AKP-Nr-1 on S. frugiperda was visualized, using a light and electron microscopy at the host-pathogen interface. Microscopic studies revealed that all the body parts of larval instars were completely overgrown by white mycelial threads of M. rileyi, except the head capsule, thoracic shield, setae, and crotchets. The cadavers of larval instars of S. frugiperda turned green on sporulation and mummified with progress in infection. In vitro pathogenicity tests revealed the potential of AKP-Nr-1 strain of M. rileyi in management of S. frugiperda. Short conclusion The results indicated the potential of M. rileyi AKP-Nr-1 as biocontrol agent for management of the fall armyworm. This AKP-Nr-1 strain of M. rileyi needs further evaluation under field conditions to evaluate its efficacy against S. frugiperda and its effects on other hosts.