{"title":"人白细胞低聚糖和糖蛋白特异性唾液酸酶的分离和鉴定。","authors":"R Schauer, M Wember, H Tschesche","doi":"10.1515/bchm2.1984.365.1.419","DOIUrl":null,"url":null,"abstract":"<p><p>A sialidase was solubilized with the aid of Triton X-100 from the insoluble material of a leucocyte homogenate. The enzyme was purified almost to homogeneity by chromatography on Sephadex G-75, equine submandibular gland mucin bound to Sepharose 4B and on Sephacryl S-200. The purification factor was 40 based on an increase of the specific enzyme activity from the Triton X-100 extract (pure enzyme: 40 mU/mg protein). Isolation of the active enzyme required the presence of a proteinase inhibitor. The sialidase is monomeric and has an average molecular mass of 48500 Da, a pH optimum of 4.6, hydrolyses preferably glycoprotein (fetuin) and sialyllactose, is activated by Ca2 and inhibited by N-acetyl-2,3-dehydro-2- deoxyneuraminic acid ( Neu5Ac2en ), Hg2 and N-(4-nitrophenyl) oxamic acid. The relatively stable enzyme shows only low activity with gangliosides and no activity with 4-O-acetylated sialic acid bound glycosidically.</p>","PeriodicalId":13015,"journal":{"name":"Hoppe-Seyler's Zeitschrift fur physiologische Chemie","volume":"365 4","pages":"419-26"},"PeriodicalIF":0.0000,"publicationDate":"1984-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1515/bchm2.1984.365.1.419","citationCount":"22","resultStr":"{\"title\":\"Isolation and characterization of an oligosaccharide- and glycoprotein-specific sialidase from human leucocytes.\",\"authors\":\"R Schauer, M Wember, H Tschesche\",\"doi\":\"10.1515/bchm2.1984.365.1.419\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>A sialidase was solubilized with the aid of Triton X-100 from the insoluble material of a leucocyte homogenate. The enzyme was purified almost to homogeneity by chromatography on Sephadex G-75, equine submandibular gland mucin bound to Sepharose 4B and on Sephacryl S-200. The purification factor was 40 based on an increase of the specific enzyme activity from the Triton X-100 extract (pure enzyme: 40 mU/mg protein). Isolation of the active enzyme required the presence of a proteinase inhibitor. The sialidase is monomeric and has an average molecular mass of 48500 Da, a pH optimum of 4.6, hydrolyses preferably glycoprotein (fetuin) and sialyllactose, is activated by Ca2 and inhibited by N-acetyl-2,3-dehydro-2- deoxyneuraminic acid ( Neu5Ac2en ), Hg2 and N-(4-nitrophenyl) oxamic acid. The relatively stable enzyme shows only low activity with gangliosides and no activity with 4-O-acetylated sialic acid bound glycosidically.</p>\",\"PeriodicalId\":13015,\"journal\":{\"name\":\"Hoppe-Seyler's Zeitschrift fur physiologische Chemie\",\"volume\":\"365 4\",\"pages\":\"419-26\"},\"PeriodicalIF\":0.0000,\"publicationDate\":\"1984-04-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://sci-hub-pdf.com/10.1515/bchm2.1984.365.1.419\",\"citationCount\":\"22\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Hoppe-Seyler's Zeitschrift fur physiologische Chemie\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://doi.org/10.1515/bchm2.1984.365.1.419\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Hoppe-Seyler's Zeitschrift fur physiologische Chemie","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.1515/bchm2.1984.365.1.419","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 22
摘要
用Triton X-100从白细胞匀浆的不溶性物质中溶解唾液酸酶。通过Sephadex G-75、马颌下腺黏液(与Sepharose 4B结合)和Sephacryl S-200的层析纯化,酶的纯度几乎一致。基于Triton X-100提取物(纯酶:40 mU/mg蛋白)的比酶活性的增加,纯化因子为40。活性酶的分离需要蛋白酶抑制剂的存在。唾液酸酶为单体,平均分子质量为48500 Da, pH最适为4.6,水解糖蛋白(fetuin)和唾液酸乳糖,受Ca2激活,受N-乙酰-2,3-脱氢-2-脱氧神经氨酸(Neu5Ac2en)、Hg2和N-(4-硝基苯基)肟酸抑制。相对稳定的酶对神经节苷的活性较低,对糖苷结合的4- o -乙酰化唾液酸无活性。
Isolation and characterization of an oligosaccharide- and glycoprotein-specific sialidase from human leucocytes.
A sialidase was solubilized with the aid of Triton X-100 from the insoluble material of a leucocyte homogenate. The enzyme was purified almost to homogeneity by chromatography on Sephadex G-75, equine submandibular gland mucin bound to Sepharose 4B and on Sephacryl S-200. The purification factor was 40 based on an increase of the specific enzyme activity from the Triton X-100 extract (pure enzyme: 40 mU/mg protein). Isolation of the active enzyme required the presence of a proteinase inhibitor. The sialidase is monomeric and has an average molecular mass of 48500 Da, a pH optimum of 4.6, hydrolyses preferably glycoprotein (fetuin) and sialyllactose, is activated by Ca2 and inhibited by N-acetyl-2,3-dehydro-2- deoxyneuraminic acid ( Neu5Ac2en ), Hg2 and N-(4-nitrophenyl) oxamic acid. The relatively stable enzyme shows only low activity with gangliosides and no activity with 4-O-acetylated sialic acid bound glycosidically.