1) using 0.3 mM NADPH and 1 mM substrate in the reduction sense, or in 100 mM Glycine-KOH buffer
(pH 10.3) using 0.3 mM NADP+ and 10 mM substrate (except for Octanol where 1 mM was used, and for 2-Chlorobenzyl alcohol and 4-Chlorobenzyl alcohol where 3 mM were used) for the oxidation sense. The specific activity towards 3,4-Dimethoxybenzaldehyde (5.1 μmol·min-1·mg-1) and to 3,4-Dimethoxybenzyl alcohol (2.0 μmol·min-1·mg-1) were taken as 100% for the reduction and oxidation reactions, {Selleck Anti-diabetic Compound Library|Selleck Antidiabetic Compound Library|Selleck Anti-diabetic Compound Library|Selleck Antidiabetic Compound Library|Selleckchem Anti-diabetic Compound Library|Selleckchem Antidiabetic Compound Library|Selleckchem Anti-diabetic Compound Library|Selleckchem Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|buy Anti-diabetic Compound Library|Anti-diabetic Compound Library ic50|Anti-diabetic Compound Library price|Anti-diabetic Compound Library cost|Anti-diabetic Compound Library solubility dmso|Anti-diabetic Compound Library purchase|Anti-diabetic Compound Library manufacturer|Anti-diabetic Compound Library research buy|Anti-diabetic Compound Library order|Anti-diabetic Compound Library mouse|Anti-diabetic Compound Library chemical structure|Anti-diabetic Compound Library mw|Anti-diabetic Compound Library molecular weight|Anti-diabetic Compound Library datasheet|Anti-diabetic Compound Library supplier|Anti-diabetic Compound Library in vitro|Anti-diabetic Compound Library cell line|Anti-diabetic Compound Library concentration|Anti-diabetic Compound Library nmr|Anti-diabetic Compound Library in vivo|Anti-diabetic Compound Library clinical trial|Anti-diabetic Compound Library cell assay|Anti-diabetic Compound Library screening|Anti-diabetic Compound Library high throughput|buy Antidiabetic Compound Library|Antidiabetic Compound Library ic50|Antidiabetic Compound Library price|Antidiabetic Compound Library cost|Antidiabetic Compound Library solubility dmso|Antidiabetic Compound Library purchase|Antidiabetic Compound Library manufacturer|Antidiabetic Compound Library research buy|Antidiabetic Compound Library order|Antidiabetic Compound Library chemical structure|Antidiabetic Compound Library datasheet|Antidiabetic Compound Library supplier|Antidiabetic Compound Library in vitro|Antidiabetic Compound Library cell line|Antidiabetic Compound Library concentration|Antidiabetic Compound Library clinical trial|Antidiabetic Compound Library cell assay|Antidiabetic Compound Library screening|Antidiabetic Compound Library high throughput|Anti-diabetic Compound high throughput screening| respectively (Table 1). The kinetic parameters K M , k cat and K i for aldehyde and alcohol substrates (Table 2) were computed by fitting initial reaction rates, measured as a function of substrate concentration, to the Michaelis-Menten equation (Equation 1) or, when substrate inhibition was observed, to the uncompetitive substrate inhibition equation (Equation 2) with the non-linear regression Enzyme Kinetics 1.3 module of the SigmaPlot 11.0 package (Systat Software, IL, USA): (1) (2) where V represents the reaction rate, V max is the limiting reaction rate, S is the substrate concentration, K M is the Michaelis constant and K i is
the substrate inhibition constant. The catalytic constant k cat of the enzyme for the different substrates was derived from . The total enzyme concentration [E] selleck products was evaluated using a protein molecular mass of 74.2 kDa. The enzyme kinetic parameters for NAD(P)H and NAD(P)+ + were determined with 0.2 mM 3,4-Dimethoxybenzaldehyde and 10 mM 3,4-Dimethoxybenzyl alcohol, respectively. Results are the mean ± SEM from at least three separate experiments. Authors’ contribution DDY participated in the design of the study, carried out the experimental
work, participated in the interpretation of the results and drafted the manuscript. JMF participated in the design and coordination of this study and helped to revise the manuscript. GMdB conceived and designed the study, coordinated the experiments, Baricitinib interpreted the results and revised the manuscript for important intellectual content. All authors read and approved the final manuscript. Acknowledgements We are very grateful to Jean-Luc PARROU and Emmanuelle TREVISIOL for scientific support and to Marie-Ange TESTE and Pierre ESCALIER for technical assistance. Dong- Dong YANG holds a Ph. D. grant from the China Scholarship Council. This work was supported in part by Region Midi Pyrénées (France) under Grant No. 09005247 and was carried out in the frame of COST Action FA0907 BIOFLAVOUR ( http://www.bioflavour.insa- toulouse.fr) under the EU’s Seventh Framework Programme for Research (FP7). References 1. Boerjan W, Ralph J, Baucher M: Lignin biosynthesis. Annu Rev Plant Biol 2003, 54:519–546.PubMedCrossRef 2.