Enzymic Properties of Purified Myrosinase from Lepidium sativum Seedlings

Author:

Durham Paul L.,Poulton Jonathan E.

Abstract

Abstract To establish the substrate specificity of the thioglucoside glucohydrolase myrosinase (EC 3.2.3.1), this enzyme was purified to homogeneity from light-grown cress (Lepidium sativum L.) seedlings by Sephadex gel filtration. Red Dye and anion exchange (FPLC Mono Q) chro­matography, and preparative isoelectric focusing. Hydrolytic activity was shown toward only 4 of the 29 synthetic and natural O-and S-glycosides tested. Highest activity was displayed with the endogenous glucosinolates benzylglucosinolate (Km, 295 μM) and sinigrin (Km, 300 μM) at an optimum pH of 5.5 in sodium citrate buffer. The synthetic glycosides PNPG (Km, 2.0 mM) and ONPG were poorer substrates at an optimum pH of 6.5 in potassium phos­phate buffer. The enzyme was inactive with all other nitrophenyl glycosides tested including PNP-a-D-glucoside and PNP-thio-β-D-glucoside, suggesting a requirement for O-β-D-glucose as the glycone moiety within these substrates. PNPG hydrolysis was stimulated 2.6-fold by ascorbate (1 mM). The enzyme exhibited no metal ion requirement and was strongly inhibited by lead nitrate, mercury chloride, and ferric chloride at 1 mM concentration. The metal chela­tors DIECA , EDTA , o-phenanthroline, and 2,2′-dipyridyl were not inhibitory, but the thiol reagents PCMS, PCMB, and N-ethylmaleimide (at 1mM) caused 50 -80% inhibition of enzyme activity.

Publisher

Walter de Gruyter GmbH

Subject

General Biochemistry, Genetics and Molecular Biology

同舟云学术

1.学者识别学者识别

2.学术分析学术分析

3.人才评估人才评估

"同舟云学术"是以全球学者为主线,采集、加工和组织学术论文而形成的新型学术文献查询和分析系统,可以对全球学者进行文献检索和人才价值评估。用户可以通过关注某些学科领域的顶尖人物而持续追踪该领域的学科进展和研究前沿。经过近期的数据扩容,当前同舟云学术共收录了国内外主流学术期刊6万余种,收集的期刊论文及会议论文总量共计约1.5亿篇,并以每天添加12000余篇中外论文的速度递增。我们也可以为用户提供个性化、定制化的学者数据。欢迎来电咨询!咨询电话:010-8811{复制后删除}0370

www.globalauthorid.com

TOP

Copyright © 2019-2024 北京同舟云网络信息技术有限公司
京公网安备11010802033243号  京ICP备18003416号-3