Purification and characterization of NAD(P)H quinone reductase from the latex of Hevea brasiliensis Mull.-Arg. (Euphorbiaceae)

NAD(P)H quinone reductase [NAD(P)H-QR] present in the latex of Hevea brasiliensis Mull-Arg. (Euphorbiaceae) was purified to homogeniety from the B-serum fraction obtained by freeze-thawing of the bottom fraction of ultracentrifuged fresh latex. The purification protocol involved acetone fractionation, heat treatment, ion exchange chromatography and affinity chromatography. The M-r determined by SDS-PAGE for the protein subunit was 21 kDa, and the molecular mass of the native enzyme estimated by get filtration was 83 kDa, indicating that the native enzyme is a homotetramer. The enzyme showed pH stability over a range of 6 to at least 10 (with an optimum at pH 8) and thermal stability up to 80degreesC. High NAD(P)H-QR activity (70%) was still retained after 10 h of preincubation at 80degreesC. A comparable substrate specificity for this enzyme was observed among menadione, p-benzoquinone, juglone, and plumbagid, with only duroquinone generating a lower activity. Positive correlations between latex NAD(P)H-QR activity and rubber yield per tapping [fresh latex (r=0.89, P<0.01), dry rubber (r=0.81, P<0.01)] together with flow time (r=0.85, P<0.01) indicated that enzyme activity could possibly be used as a marker to predict the yield potential of selected clones. (C) 2002 Elsevier Science Ltd. All rights reserved.