Location: Healthy Processed Foods ResearchTitle: Purification and characterization of a hexanol-degrading enzyme extracted from apple) Author
|Pan, Zhongli - John|
Submitted to: Journal of Agricultural and Food Chemistry
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/14/2012
Publication Date: 2/14/2012
Citation: Zhu, J., Shi, J., Pan, Z. 2012. Purification and characterization of a hexanol-degrading enzyme extracted from apple. Journal of Agricultural and Food Chemistry. 60:3246-3252. Interpretive Summary: Alcohols with more than two carbon atoms are commonly called higher or fusel alcohols. Higher alcohols are formed by yeast metabolism from amino acids and sugars and therefore are normal constituents naturally found in alcoholic beverages. An excessive concentration of the higher alcohols could distort food flavor and cause damage to the human nervous system. This research focused on developing a hexanol-degrading enzyme which was isolated from apple and showed high specificity and activity towards higher alcohols but not towards ethanol. In order to understand the biochemical characteristics of this enzyme, this study investigated the purification procedures, substrate specificity, optimum pH and temperature for activity and stability, metal ion effects, inhibitor and cofactor effects with hexanol as a substrate. The optimum conditions were pH 4.0 and 30°C for enzyme activity and pH 3.0-4.0 and temperatures below 40°C for enzyme stability.
Technical Abstract: An enzyme having activity towards n-hexanol was purified from apple and its biochemical characteristics were analyzed. The purification steps consisted of sedimentation with ammonium sulfate, DEAE Sepharose Fast Flow ion exchange chromatography and Sephadex G-100 column. The obtained enzyme had a yield of 16.00% with a specific activity of 18879.20U/mg protein and overall purification of 142.77-fold. The enzyme showed activity to isoamylol, 1-propanol, hexanol and isobutanol, but not towards methanol and ethanol. With n-hexanol as a substrate, the optimum conditions were pH 4.0 and 30°C for enzyme activity and pH 3.0-4.0 and temperatures below 40oC for enzyme stability. The enzyme activity was increased significantly by adding L-cysteine and Fe2+ at all tested concentrations and slightly by Zn2+ at a high concentration, but decreased by additions of EDTA, Ga2+, K+, Mg2+, sodium dodecyl sulfate(SDS), Sodium cholesteryl sulfate(SAS), dithiothreitol(DTT) and glutathione(GSH). The enzyme activities towards n-hexanol and n-hexanal were increased by NADH but decreased by NAD+, in contrast to a decrease towards n-hexane by both addition of NAD+ and NADH.