荧光显微法测定血浆n -乙酰- α - d -半乳糖胺酶及其一些特性的研究。

Enzyme & protein Pub Date : 1996-01-01 DOI:10.1159/000468637
W R Den Tandt, S Scharpé
{"title":"荧光显微法测定血浆n -乙酰- α - d -半乳糖胺酶及其一些特性的研究。","authors":"W R Den Tandt,&nbsp;S Scharpé","doi":"10.1159/000468637","DOIUrl":null,"url":null,"abstract":"<p><p>We have studied some characteristics of N-acetyl-alpha-D-galactosaminidase in human plasma using a sensitive and very simple fluorimetric (single tube incubation/fixation) micromethod. The enzyme has a pH optimum at 4.5, is linear on incubation during at least 6 h, is protected from inactivation at room temperature by acidification, and is stable on freezing (about 85% residual activity after 1 year at -20 degrees C). Enzyme kinetics indicate that the affinity for the substrate is low (K(m) value 7 mmol/l). By studying about 10 possible effectors, no activation was found by detergents. As expected, the colorigenic p-nitrophenyl derivative substrate, N-acetylgalactosamine and galactose are low-affinity inhibitors. A histogram of 108 control samples showed a unimodal distribution pattern with a slight bias to the right. No pseudodeficients were found on analysis of 220 control plasma samples. Patients with alpha-galactosaminidosis had residual activity between 0.7 and 2.1%. In patients with 17 different lysosomal storage diseases, no increase was found except in mucolipidosis II and III. The main advantages of the method are its simplicity sensitivity, short incubation time requirement and economy in substrate consumption. The method can be used either for screening or diagnostic purposes of genetic N-acetyl-alpha-D-galactosaminidase deficiency.</p>","PeriodicalId":11854,"journal":{"name":"Enzyme & protein","volume":null,"pages":null},"PeriodicalIF":0.0000,"publicationDate":"1996-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1159/000468637","citationCount":"3","resultStr":"{\"title\":\"Micromethod for the fluorimetric determination of plasma N-acetyl-alpha-D-galactosaminidase and study of some of its characteristics.\",\"authors\":\"W R Den Tandt,&nbsp;S Scharpé\",\"doi\":\"10.1159/000468637\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>We have studied some characteristics of N-acetyl-alpha-D-galactosaminidase in human plasma using a sensitive and very simple fluorimetric (single tube incubation/fixation) micromethod. The enzyme has a pH optimum at 4.5, is linear on incubation during at least 6 h, is protected from inactivation at room temperature by acidification, and is stable on freezing (about 85% residual activity after 1 year at -20 degrees C). Enzyme kinetics indicate that the affinity for the substrate is low (K(m) value 7 mmol/l). By studying about 10 possible effectors, no activation was found by detergents. As expected, the colorigenic p-nitrophenyl derivative substrate, N-acetylgalactosamine and galactose are low-affinity inhibitors. A histogram of 108 control samples showed a unimodal distribution pattern with a slight bias to the right. No pseudodeficients were found on analysis of 220 control plasma samples. Patients with alpha-galactosaminidosis had residual activity between 0.7 and 2.1%. In patients with 17 different lysosomal storage diseases, no increase was found except in mucolipidosis II and III. The main advantages of the method are its simplicity sensitivity, short incubation time requirement and economy in substrate consumption. The method can be used either for screening or diagnostic purposes of genetic N-acetyl-alpha-D-galactosaminidase deficiency.</p>\",\"PeriodicalId\":11854,\"journal\":{\"name\":\"Enzyme & protein\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":0.0000,\"publicationDate\":\"1996-01-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://sci-hub-pdf.com/10.1159/000468637\",\"citationCount\":\"3\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Enzyme & protein\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://doi.org/10.1159/000468637\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Enzyme & protein","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.1159/000468637","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 3

摘要

我们研究了人血浆中n -乙酰- α - d -半乳糖胺酶的一些特性,采用了一种灵敏且非常简单的荧光法(单管孵育/固定)。该酶的最佳pH值为4.5,在孵育至少6小时内呈线性,在室温下通过酸化防止失活,并且在冷冻时稳定(在-20℃下1年后约有85%的剩余活性)。酶动力学表明,该酶对底物的亲和力较低(K(m)值为7 mmol/l)。通过研究大约10种可能的效应剂,没有发现洗涤剂的活化作用。正如预期的那样,色素原对硝基苯衍生物底物、n -乙酰半乳糖胺和半乳糖是低亲和力抑制剂。108个对照样本的直方图显示单峰分布模式,有轻微的右偏。220份对照血浆样品分析未发现假缺陷。α -半乳糖胺中毒患者的残留活度在0.7 ~ 2.1%之间。在17种不同溶酶体贮积症患者中,除粘脂病II型和III型外,均未见升高。该方法的主要优点是简单、灵敏、孵育时间短、底物消耗少。该方法可用于遗传n -乙酰- α - d -半乳糖胺酶缺乏症的筛选或诊断目的。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Micromethod for the fluorimetric determination of plasma N-acetyl-alpha-D-galactosaminidase and study of some of its characteristics.

We have studied some characteristics of N-acetyl-alpha-D-galactosaminidase in human plasma using a sensitive and very simple fluorimetric (single tube incubation/fixation) micromethod. The enzyme has a pH optimum at 4.5, is linear on incubation during at least 6 h, is protected from inactivation at room temperature by acidification, and is stable on freezing (about 85% residual activity after 1 year at -20 degrees C). Enzyme kinetics indicate that the affinity for the substrate is low (K(m) value 7 mmol/l). By studying about 10 possible effectors, no activation was found by detergents. As expected, the colorigenic p-nitrophenyl derivative substrate, N-acetylgalactosamine and galactose are low-affinity inhibitors. A histogram of 108 control samples showed a unimodal distribution pattern with a slight bias to the right. No pseudodeficients were found on analysis of 220 control plasma samples. Patients with alpha-galactosaminidosis had residual activity between 0.7 and 2.1%. In patients with 17 different lysosomal storage diseases, no increase was found except in mucolipidosis II and III. The main advantages of the method are its simplicity sensitivity, short incubation time requirement and economy in substrate consumption. The method can be used either for screening or diagnostic purposes of genetic N-acetyl-alpha-D-galactosaminidase deficiency.

求助全文
通过发布文献求助,成功后即可免费获取论文全文。 去求助
来源期刊
自引率
0.00%
发文量
0
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
联系我们:info@booksci.cn Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术官方微信