BIOCHEMICAL BASIS OF SULFATASE ACTIVITY

硫酸酯酶活性的生物化学基础

• 批准号: 3324524
• 负责人: ALLEN L HORWITZ
• 金额: $ 17.55万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-05-01 至 1993-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3324524
• 关键词: antibody; arylsulfatases; complementary DNA; electrofocusing; enzyme biosynthesis; enzyme structure; fibroblasts; genetic transcription; genetic translation; human tissue; immunoprecipitation; inborn metabolism disorder; laboratory mouse; laboratory rabbit; laboratory rat; messenger RNA; mucopolysaccharidosis type II; mucopolysaccharidosis type VI; oligosaccharides; protein sequence; regulatory gene; sulfatases; tissue /cell culture; transfection

Mammalian sulfatases are a diverse group of enzymes which hydrolize steroid, mucopolysaccharide and glycolipid sulfates. The sulfatase deficiency disorders include a number of lysosomal storage diseases, and are a significant cause of mental retardation and/or physical deformities. Multiple sulfatase deficiency (MSD) is a unique genetic disorder resulting from deficiencies of at least seven distinct sulfatases and appears to result from increased degradation of cellular sulfatases. These investigations are aimed at the definition of various controls of sulfatase activity and causes of defects at various levels, including transcription, translation and post-translational processes. The elucidation of the defect in MSD is likely to demonstrate new levels of control of cellular hydrolases. Antibody and cDNA probes for steroid sulfatase, iduronate sulfatase (Id-S) and arylsulfatase B (ASB) will be used to study the various levels of control of study the molecular basis of the genetic variation of ASB activity in mouse strains with high or low ASB activity. Synthesis and processing of ASB in these mice using immunoprecipitation and pulse-chase labeling will also be studied to detect the effect of regulatory genes on mouse ASB. cDNA clones and antibodies for human ASB and human Id-S will be isolated and used to characterize the mRNA and newly synthesized polypeptides in ASB deficient Maroteaux-Lamy syndrome and Id-S deficient Hunter disease. These probes plus those for steroid sulfatase will be used to characterize the synthesis and processing of sulfatase in MSD. cDNA for sulfatase will be transfected into MSD and normal cells, and the translation and processing of the transfected STS will be followed. The subcellular compartmentalization and degradation of microinjected STS from normal and MSD cells will be evaluated. Immunoprecipitated labeled sulfatases from normal and MSD cells will be compared by isoelectric focussing, peptide mapping, oligosaccharide content and subcellular location. The role of cellular stabilization factors for sulfatases will be investigated. The route and location of post-translational processing and the role of the Golgi complex in sulfatase processing, including degradation, will be determined. The understanding of all these processes will be important in devising strategies for the treatment of enzyme deficiency diseases.

哺乳动物硫酸酯酶是一组不同的酶， 氢化类固醇、粘多糖和糖脂硫酸盐。 硫酸酯酶缺乏症包括许多溶酶体损伤。 储存疾病，是智力迟钝的重要原因 和/或身体畸形。 多发性硫酸酯酶缺乏症 是一种独特的遗传性疾病， 七种不同的硫酸酯酶，似乎是由于增加 细胞硫酸酯酶的降解。 这些调查旨在 硫酸酯酶活性的各种控制的定义， 各种层次的缺陷的原因，包括转录， 翻译和翻译后过程。 阐明 默沙东的缺陷可能会显示出新的控制水平， 细胞水解酶。 类固醇硫酸酯酶艾杜糖醛酸的抗体和cDNA探针 硫酸酯酶（Id-S）和芳基硫酸酯酶B（AS B）将用于研究 各级控制研究的分子基础 高脂血症小鼠ASB活性的遗传变异 ASB活性低。 ASB在这些小鼠体内的合成和加工 使用免疫沉淀和脉冲追踪标记， 研究了调节基因对小鼠ASB的影响。 人ASB和人Id-S的cDNA克隆和抗体将 分离并用于表征mRNA， ASB缺陷型Maroteaux-Lamy中的合成多肽 综合征和Id-S缺陷型亨特氏病。 这些探针加上 类固醇硫酸酯酶的那些将用于表征 MSD中硫酸酯酶的合成和加工。 硫酸酯酶cDNA 将其转染到MSD和正常细胞中， 并随后处理转染的STS。 的 亚细胞区室化和降解 将评价来自正常和MSD细胞的微注射STS。 来自正常和MSD细胞的免疫沉淀标记的硫酸酯酶 将通过等电聚焦，肽图谱， 寡糖含量和亚细胞位置。 的作用 将研究硫酸酯酶的细胞稳定因子。 翻译后加工的途径和位置以及翻译后加工的机制 高尔基复合体在硫酸酯酶加工中的作用，包括 退化，将被确定。 对所有这些的理解 在制定战略方面， 酶缺乏症的治疗。