{"title":"菠菜硝酸盐还原酶:通过亲和层析进一步纯化和去除“缺口”亚基","authors":"R.J. Fido, B.A. Notton","doi":"10.1016/0304-4211(84)90208-6","DOIUrl":null,"url":null,"abstract":"<div><p>NADH-nitrate reductase (EC 1.6.6.1) from spinach (<em>Spinacea oleracea</em> L. v. Noorman) has been purified 3400-fold by a multi-stage procedure involving streptomycin sulphate, (NH<sub>4</sub>)<sub>2</sub>SO<sub>4</sub>, hydroxylapatite, molecular seiving and two stages of affinity chromatography using blue-Sepharose and 5′ AMP-Sepharose. The enzyme has a specific activity of 51 μmol NO<sub>2</sub><sup>−</sup> produced min<sup>−1</sup> mg<sup>−1</sup> and a functional haem with extinction coefficients (mM) of 127 at 412 nm (oxidized) and 172 at 423 (NADH-reduced). Gel electrophoresis indicates two sub-units of approx. 110 000 and 120 000.</p></div>","PeriodicalId":20221,"journal":{"name":"Plant Science Letters","volume":null,"pages":null},"PeriodicalIF":0.0000,"publicationDate":"1984-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/0304-4211(84)90208-6","citationCount":"23","resultStr":"{\"title\":\"Spinach nitrate reductase: Further purification and removal of ‘nicked’ sub-units by affinity chromatography\",\"authors\":\"R.J. Fido, B.A. Notton\",\"doi\":\"10.1016/0304-4211(84)90208-6\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><p>NADH-nitrate reductase (EC 1.6.6.1) from spinach (<em>Spinacea oleracea</em> L. v. Noorman) has been purified 3400-fold by a multi-stage procedure involving streptomycin sulphate, (NH<sub>4</sub>)<sub>2</sub>SO<sub>4</sub>, hydroxylapatite, molecular seiving and two stages of affinity chromatography using blue-Sepharose and 5′ AMP-Sepharose. The enzyme has a specific activity of 51 μmol NO<sub>2</sub><sup>−</sup> produced min<sup>−1</sup> mg<sup>−1</sup> and a functional haem with extinction coefficients (mM) of 127 at 412 nm (oxidized) and 172 at 423 (NADH-reduced). Gel electrophoresis indicates two sub-units of approx. 110 000 and 120 000.</p></div>\",\"PeriodicalId\":20221,\"journal\":{\"name\":\"Plant Science Letters\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":0.0000,\"publicationDate\":\"1984-11-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://sci-hub-pdf.com/10.1016/0304-4211(84)90208-6\",\"citationCount\":\"23\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Plant Science Letters\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/0304421184902086\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Plant Science Letters","FirstCategoryId":"1085","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/0304421184902086","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 23
摘要
从菠菜(Spinacea oleracea L. v. Noorman)中纯化出nadh -硝酸盐还原酶(EC 1.6.6.1)，该酶采用多阶段程序，包括硫酸链霉素、(NH4)2SO4、羟基磷灰石、分子分离以及使用blue-Sepharose和5 ' AMP-Sepharose的两阶段亲和层析，纯化了3400倍。该酶产生的NO2−比活性为51 μmol min−1 mg−1，在412 nm(氧化)处消光系数为127，在423 nm (nadh还原)处消光系数为172。凝胶电泳显示两个亚基约。11万和12万。
Spinach nitrate reductase: Further purification and removal of ‘nicked’ sub-units by affinity chromatography
NADH-nitrate reductase (EC 1.6.6.1) from spinach (Spinacea oleracea L. v. Noorman) has been purified 3400-fold by a multi-stage procedure involving streptomycin sulphate, (NH4)2SO4, hydroxylapatite, molecular seiving and two stages of affinity chromatography using blue-Sepharose and 5′ AMP-Sepharose. The enzyme has a specific activity of 51 μmol NO2− produced min−1 mg−1 and a functional haem with extinction coefficients (mM) of 127 at 412 nm (oxidized) and 172 at 423 (NADH-reduced). Gel electrophoresis indicates two sub-units of approx. 110 000 and 120 000.
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