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d-pantolactone is an intermediate in the synthesis of d-pantothenic acid, which is known as vitamin B5. The commercial synthesis of d-pantolactone is carried out through the selective resolution of dl-pantolactone catalyzed by lactone hydrolase. In contrast to a kinetic resolution approach, the deracemization of dl-pantolactone is a simpler, greener, and more sustainable way to obtain d-pantolactone with high optical purity. Herein, an efficient three-enzyme cascade was developed for the deracemization of dl-pantolactone, using l-pantolactone dehydrogenase from Amycolatopsis methanolica (AmeLPLDH), conjugated polyketone reductase from Zygosaccharomyces parabailii (ZpaCPR), and glucose dehydrogenase from Bacillus subtilis (BsGDH). The AmeLPLDH was used to catalyze the dehydrogenated l-pantolactone into ketopantolactone; the ZpaCPR was used to further catalyze the ketopantolactone into d-pantolactone; and glucose dehydrogenase together with glucose fulfilled the function of coenzyme regeneration. All three enzymes were co-expressed in E. coli strain BL21(DE3), which served as the whole-cell biocatalyst. Under optimized conditions, 36 h deracemization of 1.25 M dl-pantolactone d-pantolactone led to an e.e.p value of 98.6%, corresponding to productivity of 107.7 g/(l·d).

Glycolate oxidase is a peroxisomal flavoprotein catalyzing the oxidation of glycolate to glyoxylate and plays crucial metabolic roles in green algae, plants, and animals. It could serve as a biocatalyst for enzymatic production of glyoxylate, a fine chemical with a wide variety of applications in perfumery, flavor, and the pharmaceutical and agrochemical industries. However, the low catalytic activity of native glycolate oxidase and low levels of active enzyme in heterologous expression limit its practical use in industrial biocatalysis. Herein, the glycolate oxidase from Chlamydomonas reinhardtii (CreGO) was selected through phylogenetic tree analysis, and its low level of soluble expression in E. coli BL21(DE3) was improved through the use of the glutathione thioltransferase (GST), the choice of the vector pET22b and the optimization of induction conditions. The semi-rational design of the fusion enzyme GST-Gly-Ser-Gly-CreGO led to the superior variant GST-Gly-Ser-Gly-CreGO-Y27S/V111G/V212R with the kcat/Km value of 29.2 s−1·mM−1, which was six times higher than that of the wild type. In contrast to GST-Gly-Ser-Gly-CreGO, 5 mg/mL of crude enzyme GST-Gly-Ser-Gly-CreGO-Y27S/V111G/V212R together with 25 μg/mL of catalase catalyzed the oxidation of 300 mM of methyl glycolate for 8 h, increasing the yield from 50.4 to 93.5%.

Abstract Background α,β-Unsaturated aldehydes are widely used in the organic synthesis of fine chemicals for application in products such as flavoring agents, fragrances and pharmaceuticals. In the selective oxidation of α,β-unsaturated alcohols to the corresponding α,β-unsaturated aldehydes, it remains challenging to overcome poor selectivity, overoxidation and a low atom efficiency in chemical routes. Results An E. coli strain coexpressing the NADP+-specific alcohol dehydrogenase YsADH and the oxygen-dependent NADPH oxidase TkNOX was constructed; these components enabled the NADP+ regeneration and catalyzed the oxidation of 100 mM 3-methyl-2-buten-1-ol to 3-methyl-2-butenal with a yield of 21.3%. The oxygen supply was strengthened by introducing the hemoglobin protein VsHGB into recombinant E. coli cells and replacing the atmosphere of the reactor with pure oxygen, which increased the yield to 51.3%. To further improve catalytic performance, the E. coli cells expressing the multifunctional fusion enzyme YsADH-(GSG)-TkNOX-(GSG)-VsHGB were generated, which completely converted 250 mM 3-methyl-2-buten-1-ol to 3-methyl-2-butenal after 8 h of whole-cell oxidation. The reaction conditions for the cascade biocatalysis were optimized, in which supplementation with 0.2 mM FAD and 0.4 mM NADP+ was essential for maintaining high catalytic activity. Finally, the established whole-cell system could serve as a platform for the synthesis of valuable α,β-unsaturated aldehydes through the selective oxidation of various α,β-unsaturated alcohols. Conclusions The construction of a strain expressing the fusion enzyme YsADH-(GSG)-TkNOX-(GSG)-VsHGB achieved efficient NADP+ regeneration and the selective oxidation of various α,β-unsaturated alcohols to the corresponding α,β-unsaturated aldehydes. Among the available redox enzymes, the fusion enzyme YsADH-(GSG)-TkNOX-(GSG)-VsHGB has become the most recent successful example to improve catalytic performance in comparison with its separate components.

The cascade catalysis of old yellow enzyme, alcohol dehydrogenase and glucose dehydrogenase has become a promising approach for one pot, two-step reduction of (E/Z)-citral to (S)-citronellol, serving as a chiral alcohol with rose fragrance. During the multi-enzymatic cascade catalysis, old yellow enzyme is responsible for the reduction of the conjugated C=C and the introduction of the chiral center, requiring high activity and (S)-enantioselectiviy. Herein, to improve the activity of the old yellow enzyme from Providencia stuartii (NemR-PS) with strict (S)-enantioselectivity, the semi-rational design on its substrate binding pocket was performed through a combination of homology modeling, molecular docking analysis, alanine scanning and iterative saturation mutagenesis. The NemR-PS variant D275G/F351A with improved activity was obtained and then purified for characterization, obeying the substrate inhibition kinetics. Compared with the wild type, the parameters Ki and Kcat/Km were increased from 39.79 mM and 2.09 s−1mM−1 to 128.50 mM and 5.01 s−1mM−1, respectively. Moreover, the variant D275G/F351A maintained strict (S)-enantioselectivity, avoiding the trade-off effect between activity and enantioselectivity. Either the enzyme NemR-PS or the variant D275G/F351A was co-expressed with alcohol dehydrogenase from Yokenella sp. WZY002 (YsADH) and glucose dehydrogenase from Bacillus megaterium (BmGDHM6). In contrast to the whole-cell biocatalyst co-expressing NemR-PS, that co-expressing the variant D275G/F351A shortened the reaction time from 36 h to 12 h in the reduction of 400 mM (E/Z)-citral. In the manner of substrate constant feeding, the accumulated product concentration reached up to 500 mM and completely eliminate the residual intermediate and by-product, suggesting the effectiveness of protein engineering and substrate engineering to improve catalytic efficiency.

The use of nanomedicines for cancer treatment has been widespread. Fullerenes have significant effects in the treatment of solid tumors. Here, we are going to study the effects of hydroxylated fullerene C60(OH)n(n = 18–22) treatment on chronic myeloid leukemia cell proliferation and investigate its toxicity. The results showed that hydroxylated fullerene C60(OH)n (n = 18–22) at low concentrations (less than 120 μM) not only had apparent toxic side effects, but also promoted the growth of K562 cells, while a high concentration of C60(OH)n had different degrees of inhibition on K562 cells. When the concentration is higher than 160 μM, the K562 cells showed morphological changes, the mitochondrial membrane potential decreased, the cell cycle was blocked in the stage of G2-phase, and cell apoptosis occurred, which may cause apoptosis, autophagy, and a variety of other damage leading to cell death. Meanwhile, it also indicated that its inhibition of solid tumors might be related to the tumor microenvironment; we verified the safety of fullerene without apparent cellular toxicity at a specific concentration.

The importance of yeast old yellow enzymes is increasingly recognized for direct asymmetric reduction of (E/Z)-citral to (R)-citronellal. As one of the most performing old yellow enzymes, the enzyme OYE3 from Saccharomyces cerevisiae S288C exhibited complementary enantioselectivity for the reduction of (E)-citral and (Z)-citral, resulting in lower e.e. value of (R)-citronellal in the reduction of (E/Z)-citral. To develop a novel approach for the direct synthesis of enantio-pure (R)-citronellal from the reduction of (E/Z)-citral, the enzyme OYE3 was firstly modified by semi-rational design to improve its (R)-enantioselectivity. The OYE3 variants W116A and S296F showed strict (R)-enantioselectivity in the reduction of (E)-citral, and significantly reversed the (S)-enantioselectivity in the reduction of (Z)-citral. Next, the double substitution of OYE3 led to the unique variant S296F/W116G, which exhibited strict (R)-enantioselectivity in the reduction of (E)-citral and (E/Z)-citral, but was not active on (Z)-citral. Relying on its capability discriminating (E)-citral and (Z)-citral, a new cascade reaction catalyzed by the OYE3 variant S296F/W116G and glucose dehydrogenase was developed, providing the enantio-pure (R)-citronellal and the retained (Z)-citral after complete reduction of (E)-citral.

Citronellol is a kind of unsaturated alcohol with rose-like smell and its (S)-enantiomer serves as an important intermediate for organic synthesis of (-)-cis-rose oxide. Chemical methods are commonly used for the synthesis of citronellol and its (S)-enantiomer, which suffers from severe reaction conditions and poor selectivity. Here, the first one-pot double reduction of (E/Z)-citral to (S)-citronellol was achieved in a multi-enzymatic cascade system: N-ethylmaleimide reductase from Providencia stuartii (NemR-PS) was selected to catalyze the selective reduction of (E/Z)-citral to (S)-citronellal, alcohol dehydrogenase from Yokenella sp. WZY002 (YsADH) performed the further reduction of (S)-citronellal to (S)-citronellol, meanwhile a variant of glucose dehydrogenase from Bacillus megaterium (BmGDHM6), together with glucose, drove efficient NADPH regeneration. The Escherichia coli strain co-expressing NemR-PS, YsADH, and BmGDHM6 was successfully constructed and used as the whole-cell catalyst. Various factors were investigated for achieving high conversion and reducing the accumulation of the intermediate (S)-citronellal and by-products. 0.4 mM NADP+ was essential for maintaining high catalytic activity, while the feeding of the cells expressing BmGDHM6 effectively eliminated the intermediate and by-products and shortened the reaction time. Under optimized conditions, the bio-transformation of 400 mM citral caused nearly complete conversion (>99.5%) to enantio-pure (S)-citronellol within 36 h, demonstrating promise for industrial application.

Methionine (Met) is an essential sulfur-containing amino acid in animals. Cereal and legume crops with limiting levels of Met represent the major food and feed sources for animals. In plants, cystathionine gamma-synthase (CGS), methionine methyltransferase (MMT) and homocysteine methyltransferase (HMT) are committing enzymes synergistically synthesizing Met through the aspartate (Asp) family pathway and the S-methylmethionine (SMM) cycle. The biological functions of CGS, MMT and HMT genes have been respectively studied, whereas their evolution patterns and their contribution to the evolution of Met biosynthetic pathway in plants are unknown. In the present study, to reveal their evolution patterns and contribution, the evolutionary relationship of CGS, MMT and HMT gene families were reconstructed. The results showed that MMTs began in the ancestor of the land plants and kept conserved during evolution, while the CGSs and HMTs had diverged. The CGS genes were divided into two branches in the angiosperms, Class 1 and Class 2, of which Class 2 only contained the grasses. However, the HMT genes diverged into Class 1 and Class 2 in all of the seed plants. Further, the gene structure analysis revealed that the CGSs, MMTs and HMTs were relatively conserved except for the CGSs in Class 2. According to the expression of CGS, HMT and MMT genes in soybeans, as well as in the database of soybean, rice and Arabidopsis, the expression patterns of the MMTs were shown to be consistently higher in leaves than in seeds. However, the expression of CGSs and HMTs had diverged, either expressed higher in leaves or seeds, or showing fluctuated expression. Additionally, the functions of HMT genes had diverged into the repair of S-adenosylmethionine and SMM catabolism during the evolution. The results indicated that the CGS and HMT genes have experienced partial subfunctionalization. Finally, given the evolution and expression of the CGS, HMT and MMT gene families, we built the evolutionary model of the Met biosynthetic pathways in plants. The model proposed that the Asp family pathway existed in all the plant lineages, while the SMM cycle began in the ancestor of land plants and then began to diverge in the ancestor of seed plants. The model suggested that the evolution of Met biosynthetic pathway is basically consistent with that of plants, which might be vital to the growth and development of different botanical lineages during evolution.

The development of novel nanoparticles as a new generation therapeutic drug platform is an active field of chemistry and cancer research. In recent years, fullerene nanoparticles have received extensive attention due to their unique physical and chemical properties. Properly modified fullerene nanoparticles have excellent biocompatibility and significant anti-tumor activity, which makes them have broad application prospects in the field of cancer therapy. Therefore, understanding the anti-tumor mechanism of fullerene nanoparticles is of great significance for the design and development of anti-tumor drugs with low toxicity and high targeting. This review has focused on various anti-tumor mechanisms of fullerene derivatives and discusses their toxicity and their distribution in organisms. Finally, the review points out some urgent problems that need solution before fullerene derivatives as a new generation of anti-tumor nano-drug platform enter clinical research.

Possessing aldehyde and carboxyl groups, glyoxylic acid and its ester derivatives serve as platform chemicals for the synthesis of vanillin, (R)-pantolactone, antibiotics or agrochemicals. Methyl glycolate is one of the by-products in the coal-to-glycol industry, and we attempted its value-added use through enzymatic oxidation of methyl glycolate to methyl glyoxylate. The cascade catalysis of glycolate oxidase from Spinacia oleracea (SoGOX), catalase from Helicobacter pylori (HpCAT) and hemoglobin from Vitreoscilla stercoraria (VsHGB) was firstly constructed, despite poor catalytic performance. To enable efficient oxidation of methyl glycolate, eight fusion enzymes of SoGOX, HpCAT and VsHGB were constructed by varying the orientation and the linker length. The fusion enzyme VsHGB-GSG-SoGOX-GGGGS-HpCAT was proved to be best, which reaction yield was 2.9 times higher than that of separated enzymes. The enzyme SoGOX was further subjected to directed evolution and site-saturation mutagenesis. The reaction yield of the resulting variant M267T/S362G was 1.9 times higher than that of the wild type. Then, the double substitution M267T/S362G was integrated with fusion expression to give the fusion enzyme VsHGB-GSG-SoGOXmut-GGGGS-HpCAT, which crude enzyme was used as biocatalyst. The use of crude enzyme virtually eliminated side reactions and simplified the preparation of biocatalysts. Under the optimized conditions, the crude enzyme VsHGB-GSG-SoGOXmut-GGGGS-HpCAT catalyzed the oxidation of 200 mM methyl glycolate for 6 h, giving a yield of 95.3%. The development of efficient fusion enzyme and the use of its crude enzyme paved the way for preparative scale application on enzymatic oxidation of methyl glycolate to methyl glyoxylate.

The gene encoding a (2R,3R)-2,3-butanediol dehydrogenase from Rhodococcus erythropolis WZ010 (ReBDH) was over-expressed in Escherichia coli and the resulting recombinant ReBDH was successfully purified by Ni-affinity chromatography. The purified ReBDH in the native form was found to exist as a monomer with a calculated subunit size of 37180, belonging to the family of the zinc-containing alcohol dehydrogenases. The enzyme was NAD(H)-specific and its optimal activity for acetoin reduction was observed at pH 6.5 and 55 °C. The optimal pH and temperature for 2,3-butanediol oxidation were pH 10 and 45 °C, respectively. The enzyme activity was inhibited by ethylenediaminetetraacetic acid (EDTA) or metal ions Al3+, Zn2+, Fe2+, Cu2+ and Ag+, while the addition of 10% (v/v) dimethyl sulfoxide (DMSO) in the reaction mixture increased the activity by 161.2%. Kinetic parameters of the enzyme showed lower Km values and higher catalytic efficiency for diacetyl and NADH in comparison to those for (2R,3R)-2,3-butanediol and NAD+. The activity of acetoin reduction was 7.7 times higher than that of (2R,3R)-2,3-butanediol oxidation when ReBDH was assayed at pH 7.0, suggesting that ReBDH-catalyzed reaction in vivo might favor (2R,3R)-2,3-butanediol formation rather than (2R,3R)-2,3-butanediol oxidation. The enzyme displayed absolute stereospecificity in the reduction of diacetyl to (2R,3R)-2,3-butanediol via (R)-acetoin, demonstrating its potential application on the synthesis of (R)-chiral alcohols.

The members of the Old Yellow Enzyme (OYE) family are capable of catalyzing the asymmetric reduction of (E/Z)-citral to (R)-citronellal—a key intermediate in the synthesis of L-menthol. The applications of OYE-mediated biotransformation are usually hampered by its insufficient enantioselectivity and low activity. Here, the (R)-enantioselectivity of Old Yellow Enzyme from Saccharomyces cerevisiae CICC1060 (OYE2y) was enhanced through protein engineering. The single mutations of OYE2y revealed that the sites R330 and P76 could act as the enantioselectivity switch of OYE2y. Site-saturation mutagenesis was conducted to generate all possible replacements for the sites R330 and P76, yielding 17 and five variants with improved (R)-enantioselectivity in the (E/Z)-citral reduction, respectively. Among them, the variants R330H and P76C partly reversed the neral derived enantioselectivity from 32.66% e.e. (S) to 71.92% e.e. (R) and 37.50% e.e. (R), respectively. The docking analysis of OYE2y and its variants revealed that the substitutions R330H and P76C enabled neral to bind with a flipped orientation in the active site and thus reverse the enantioselectivity. Remarkably, the double substitutions of R330H/P76M, P76G/R330H, or P76S/R330H further improved (R)-enantioselectivity to >99% e.e. in the reduction of (E)-citral or (E/Z)-citral. The results demonstrated that it was feasible to alter the enantioselectivity of OYEs through engineering key residue distant from active sites, e.g., R330 in OYE2y.

Unsaturated fatty acids are the main components of vegetable oils. Fatty acid desaturase 2 (FAD2) catalyzes oleic acid (OA) into linoleic acid (LA) transformations, which are essential to the profile of FAs in seeds. To further understand the roles of FAD2s in the synthesis of oil, the evolution and biocatalysis of FAD2s were comprehensively analyzed. The evolution history of the FAD2 gene family showed that most of the FAD2 genes formed monophyletic clades except in eudicots. The FAD2 genes in some eudicots diverged into constitutive and seed-specific expression clades. Notably, the biocatalysis of seed-specific or -abundant expression FAD2s in soybean, perilla, rice, and spruce revealed that their catalytic activity was strongly correlated with the total oil content of their seeds in nature. Additionally, it was found that I and Y in site 143 of GmaFAD2-1 were strictly conserved in the seed-specific and constitutive expression clades of Fabaceae, respectively. Furthermore, the site-directed mutation demonstrated that I and Y are vital to improving and reducing the activity of GmaFAD2s. Therefore, the results indicate that the activity of FAD2s in seeds might be a reference to the total oil content of seeds, and site 143 might have been specifically evolved to be required for the activity of FAD2s in some expression-diverged eudicots, especially in legumes.

The recombinant carbonyl reductase from Rhodococcus erythropolis WZ010 (ReCR) demonstrated strict (S)-stereoselectivity and catalyzed the irreversible reduction of N-Boc-3-piperidone (NBPO) to (S)-N-Boc-3-hydroxypiperidine [(S)-NBHP], a key chiral intermediate in the synthesis of ibrutinib. The NAD(H)-specific enzyme was active within broad ranges of pH and temperature and had remarkable activity in the presence of higher concentration of organic solvents. The amino acid residue at position 54 was critical for the activity and the substitution of Tyr54 to Phe significantly enhanced the catalytic efficiency of ReCR. The kcat/Km values of ReCR Y54F for NBPO, (R/S)-2-octanol, and 2-propanol were 49.17 s−1 mM−1, 56.56 s−1 mM−1, and 20.69 s−1 mM−1, respectively. In addition, the (S)-NBHP yield was as high as 95.92% when whole cells of E. coli overexpressing ReCR variant Y54F catalyzed the asymmetric reduction of 1.5 M NBPO for 12 h in the aqueous/(R/S)-2-octanol biphasic system, demonstrating the great potential of ReCR variant Y54F for practical applications.

Homocysteine methyltransferase (HMT) converts homocysteine to methionine using S-methylmethionine (SMM) or S-adenosylmethionine (SAM) as methyl donors in organisms, playing an important role in supplying methionine for the growth and the development of plants. To better understand the functions of the HMT genes in plants, we conducted a wide evolution and expression analysis of these genes. Reconstruction of the phylogenetic relationship showed that the HMT gene family was divided into Class 1 and Class 2. In Class 1, HMTs were only found in seed plants, while Class 2 presented in all land plants, which hinted that the HMT genes might have diverged in seed plants. The analysis of gene structures and selection pressures showed that they were relatively conserved during evolution. However, type I functional divergence had been detected in the HMTs. Furthermore, the expression profiles of HMTs showed their distinct expression patterns in different tissues, in which some HMTs were widely expressed in various organs, whereas the others were highly expressed in some specific organs, such as seeds or leaves. Therefore, according to our results in the evolution, functional divergence, and expression, the HMT genes might have diverged during evolution. Further analysis in the expression patterns of AthHMTs with their methyl donors suggested that the diverged HMTs might be related to supply methionine for the development of plant seeds.

The gene encoding a thermostable iron-containing alcohol dehydrogenase from Thermococcus Strain ES1 (ES1 ADH) was cloned, sequenced and expressed in Escherichia coli. The recombinant and native ES1 ADHs were purified using multistep column chromatography under anaerobic conditions. Both enzymes appeared to be homotetramers with a subunit size of 45 ± 1 kDa as revealed by SDS-PAGE, which was close to the calculated value (44.8 kDa). The recombinant ADH contained 1.0 ± 0.1 g-atom iron per subunit. Both enzymes were sensitive to oxygen with a half-life upon exposure to air of about 4 min. The recombinant enzyme exhibited a specific activity of 105 ± 2 U mg−1, which was very similar to that of the native enzyme (110 ± 3 U mg−1). The optimal pH-values for both enzymes for ethanol oxidation and acetaldehyde reduction were 10.4 and 7.0, respectively. Both enzymes also showed similar temperature-dependent activities, and catalyzed the oxidation of primary alcohols, but there was no activity towards methanol and secondary alcohols. Kinetic parameters of the enzymes showed lower Km-values for acetaldehyde and NADPH and higher Km-values for ethanol and NADP+. It is concluded that the gene encoding ES1 ADH was expressed successfully in E. coli. This is the first report of a fully active recombinant version of an iron-containing ADH from a hyperthermophile. [ABSTRACT FROM AUTHOR]

Thermotoga hypogea is an extremely thermophilic anaerobic bacterium capable of growing at 90°C. It uses carbohydrates and peptides as carbon and energy sources to produce acetate, CO2, H2, l-alanine and ethanol as end products. Alcohol dehydrogenase activity was found to be present in the soluble fraction of T. hypogea. The alcohol dehydrogenase was purified to homogeneity, which appeared to be a homodimer with a subunit molecular mass of 40 ± 1 kDa revealed by SDS-PAGE analyses. A fully active enzyme contained iron of 1.02 ± 0.06 g-atoms/subunit. It was oxygen sensitive; however, loss of enzyme activity by exposure to oxygen could be recovered by incubation with dithiothreitol and Fe2+. The enzyme was thermostable with a half-life of about 10 h at 70°C, and its catalytic activity increased along with the rise of temperature up to 95°C. Optimal pH values for production and oxidation of alcohol were 8.0 and 11.0, respectively. The enzyme had a broad specificity to use primary alcohols and aldehydes as substrates. Apparent K m values for ethanol and 1-butanol were much higher than that of acetaldehyde and butyraldehyde. It was concluded that the physiological role of this enzyme is likely to catalyze the reduction of aldehydes to alcohols. [ABSTRACT FROM AUTHOR]

An alcohol dehydrogenase (ADH) from hyperthermophilic archaeon Thermococcus guaymasensis was purified to homogeneity and was found to be a homotetramer with a subunit size of 40 ± 1 kDa. The gene encoding the enzyme was cloned and sequenced; this gene had 1,095 bp, corresponding to 365 amino acids, and showed high sequence homology to zinc-containing ADHs and L-threonine dehydrogenases with binding motifs of catalytic zinc and NADP+. Metal analyses revealed that this NADP+-dependent enzyme contained 0.9 ± 0.03 g-atoms of zinc per subunit. It was a primary-secondary ADH and exhibited a substrate preference for secondary alcohols and corresponding ketones. Particularly, the enzyme with unusual stereoselectivity catalyzed an anti-Prelog reduction of racemic (R/S)-acetoin to (2R,3R)-2,3-butanediol and meso-2,3-butanediol. The optimal pH values for the oxidation and formation of alcohols were 10.5 and 7.5, respectively. Besides being hyperthermostable, the enzyme activity increased as the temperature was elevated up to 95°C. The enzyme was active in the presence of methanol up to 40% (vol/vol) in the assay mixture. The reduction of ketones underwent high efficiency by coupling with excess isopropanol to regenerate NADPH. The kinetic parameters of the enzyme showed that the apparent Km values and catalytic efficiency for NADPH were 40 times lower and 5 times higher than those for NADP+, respectively. The physiological roles of the enzyme were proposed to be in the formation of alcohols such as ethanol or acetoin concomitant to the NADPH oxidation. [ABSTRACT FROM AUTHOR]