Production of aldehyde oxidase by Streptomyces species

Date
1995
Authors
Lee, Byungtae
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Anthony L. Pometto, III
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Food Science and Human Nutrition
Abstract

Production of aldehyde oxidase (aldehyde:O[subscript]2 oxidoreductase, EC 1.2.3.1) by Streptomyces setonii 75Vi2 and Streptomyces viridosporus T7A was characterized. For S. setonii, aldehyde oxidase was initially induced in shake-flask culture in 0.6% (w/v) yeast extract medium. Inducer (propanal) concentration was determined to be 1.6 g/L medium and two propanal additions generated the best results. Inducer addition to cells in late-log phase was essential to enzyme induction. Dissolved oxygen and pH measurements were key parameters for determination of late-log phase in 15-L and 50-L batch fermentation. Aldehyde oxidase activity was 0.02 units (one unit was defined as 0.1 [mu]mole/min/mg protein) in shake-flask and CSR cultures, and 0.05, 0.1, and 0.21 units in 5-L, 15-L, and 50-L batch fermentations, respectively. However, with propanal or trans-cinnamaldehyde (aromatic aldehyde) as inducer, aldehyde oxidase production by S. setonii was inconsistent and not reproducible;For S. viridosporus, vanillin (2 g/L medium) was used as inducer in 50-L fermentations in 0.6% (w/v) yeast extract and 1.0% (w/v) malt extract medium. Vanillin was a stable enzyme inducer and its oxidation to vanillic acid was monitored spectrophotometrically at 345 nm. Aldehyde oxidase activity was more stable in S. viridosporus than S. setonii and freeze-dried cell-free extract extended its shelf-life. Propanal oxidation to propionic acid by aldehyde oxidase was confirmed by HPLC for the 30% ammonium sulfate precipitate from a cell-free extract. Oxygraphic enzyme activity measurement did not always correlate with acid production due to interference of endogenous catalase but oxygraph was beneficial for rapid aldehyde oxidase detection;Crude cell-free extract, 45% ammonium sulfate precipitate and heat-treated (70°C) for 5 and 10 min of crude extract revealed positive aldehyde oxidase activity as shown by brown color formation on nondenaturing polyacrylamide gel electrophoresis (PAGE) zymogram with vanillin as the substrate. This same band from the nondenaturing PAGE demonstrated two broad protein bands on SDS-PAGE which corresponded to peptides of 20 and 55 kDa. These bacterial oxidases were active toward propanal, hexanal, trans-cinnamaldehyde, and vanillin, which suggests its possible use to reduce off-flavors in soy products.

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