{"title":"大豆和兔网状细胞中的脂氧合酶:失活和铁释放。","authors":"W E Höhne, N Kojima, B Thiele, S M Rapoport","doi":"","DOIUrl":null,"url":null,"abstract":"<p><p>Various inactivation methods were applied to lipoxygenases from soybean (isoenzyme 1) and rabbit reticulocytes to compare the inactivation behaviour of both enzymes and to elucidate the state of the iron which is known to be involved in the catalytic reaction of lipoxygenases: 1. Titration of the enzyme with mercury compounds shows that there are one or two SH groups responsible for the loss of activity in the presence of mercury. The SH groups seem not to be involved in the tight iron binding. 2. Inactivation by chelating agents such as o-phenanthroline or batho-phenanthroline sulfonic acid occurs only in the presence of reducing agents (mercaptoethanol and ascorbic acid). Our data support a co-oxidation mechanism. The complexation of iron by chelators is not the rate-limiting step. Both lipoxygenases show a very similar behaviour in this respect despite the fact that the reticulocyte enzyme requires the addition of trace amounts of copper ions for efficient inactivation. 3. Release of iron from the enzyme is also achieved by denaturation with guanidinium hydrochloride (Gu-HCl). In all cases, inactivation and release of iron were irreversible processes. 4. A sequence comparison for both animal and plant lipoxygenases shows strongly conserved amino acids, especially histidines and hydrophobic residues, which possibly may be involved in iron complexation and substrate binding.</p>","PeriodicalId":8948,"journal":{"name":"Biomedica biochimica acta","volume":null,"pages":null},"PeriodicalIF":0.0000,"publicationDate":"1991-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Lipoxygenases from soybeans and rabbit reticulocytes: inactivation and iron release.\",\"authors\":\"W E Höhne, N Kojima, B Thiele, S M Rapoport\",\"doi\":\"\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>Various inactivation methods were applied to lipoxygenases from soybean (isoenzyme 1) and rabbit reticulocytes to compare the inactivation behaviour of both enzymes and to elucidate the state of the iron which is known to be involved in the catalytic reaction of lipoxygenases: 1. Titration of the enzyme with mercury compounds shows that there are one or two SH groups responsible for the loss of activity in the presence of mercury. The SH groups seem not to be involved in the tight iron binding. 2. Inactivation by chelating agents such as o-phenanthroline or batho-phenanthroline sulfonic acid occurs only in the presence of reducing agents (mercaptoethanol and ascorbic acid). Our data support a co-oxidation mechanism. The complexation of iron by chelators is not the rate-limiting step. Both lipoxygenases show a very similar behaviour in this respect despite the fact that the reticulocyte enzyme requires the addition of trace amounts of copper ions for efficient inactivation. 3. Release of iron from the enzyme is also achieved by denaturation with guanidinium hydrochloride (Gu-HCl). In all cases, inactivation and release of iron were irreversible processes. 4. A sequence comparison for both animal and plant lipoxygenases shows strongly conserved amino acids, especially histidines and hydrophobic residues, which possibly may be involved in iron complexation and substrate binding.</p>\",\"PeriodicalId\":8948,\"journal\":{\"name\":\"Biomedica biochimica acta\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":0.0000,\"publicationDate\":\"1991-01-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Biomedica biochimica acta\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Biomedica biochimica acta","FirstCategoryId":"1085","ListUrlMain":"","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 0
摘要
对来自大豆(同工酶 1)和家兔网状细胞的脂氧合酶采用了各种灭活方法,以比较这两种酶的灭活行为,并阐明已知参与脂氧合酶催化反应的铁的状态:1.1. 用汞化合物滴定酶的结果表明,有一个或两个 SH 基团导致酶在汞存在下失去活性。这些 SH 基团似乎与铁的紧密结合无关。2.只有在还原剂(巯基乙醇和抗坏血酸)存在的情况下,邻菲罗啉或浴菲罗啉磺酸等螯合剂才会使酶失活。我们的数据支持共同氧化机制。螯合剂对铁的络合作用不是限速步骤。尽管网状细胞酶需要添加微量铜离子才能有效失活,但两种脂氧合酶在这方面的表现非常相似。3.3. 用盐酸胍(Gu-HCl)变性也可使铁从酶中释放出来。在所有情况下,铁的失活和释放都是不可逆的过程。4.4. 对动物和植物脂氧合酶的序列进行比较后发现,它们的氨基酸,尤其是组氨酸和疏水残基有很强的保守性,这些氨基酸可能参与了铁的复合和底物的结合。
Lipoxygenases from soybeans and rabbit reticulocytes: inactivation and iron release.
Various inactivation methods were applied to lipoxygenases from soybean (isoenzyme 1) and rabbit reticulocytes to compare the inactivation behaviour of both enzymes and to elucidate the state of the iron which is known to be involved in the catalytic reaction of lipoxygenases: 1. Titration of the enzyme with mercury compounds shows that there are one or two SH groups responsible for the loss of activity in the presence of mercury. The SH groups seem not to be involved in the tight iron binding. 2. Inactivation by chelating agents such as o-phenanthroline or batho-phenanthroline sulfonic acid occurs only in the presence of reducing agents (mercaptoethanol and ascorbic acid). Our data support a co-oxidation mechanism. The complexation of iron by chelators is not the rate-limiting step. Both lipoxygenases show a very similar behaviour in this respect despite the fact that the reticulocyte enzyme requires the addition of trace amounts of copper ions for efficient inactivation. 3. Release of iron from the enzyme is also achieved by denaturation with guanidinium hydrochloride (Gu-HCl). In all cases, inactivation and release of iron were irreversible processes. 4. A sequence comparison for both animal and plant lipoxygenases shows strongly conserved amino acids, especially histidines and hydrophobic residues, which possibly may be involved in iron complexation and substrate binding.