猫和人n -乙酰半乳糖胺4-硫酸酯酶的免疫化学特性

Brooks D.A., Gibson G.J., Hopwood J.J.
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引用次数: 11

摘要

马罗托-拉米综合征(粘多糖病VI型);MPS VI)是一种由溶酶体相关酶n -乙酰半乳糖胺4-硫酸酯酶(4-硫酸酯酶)缺乏引起的疾病。先前已经描述了人类MPS VI的猫模型,并为酶替代治疗方案的评估提供了一个系统。作为人4-硫酸酯酶在猫体内替代治疗的初步研究,我们比较了人和猫4-硫酸酯酶的免疫化学性质。通过SDS-PAGE分析,纯化猫和人4-硫酸酯酶在还原和非还原条件下的分子质量相似。然而,人类和猫的4-硫酸酯酶之间存在可检测到的构象差异,表明存在一些结构差异。在免疫结合实验中,猫4-硫酸酯酶与一组单克隆抗体反应较弱(在游离溶液中与4-硫酸酯酶相互作用),但在免疫定量实验中,同样的单克隆抗体与猫4-硫酸酯酶反应强烈,其中猫4-硫酸酯酶与多克隆抗体结合(这可能导致猫4-硫酸酯酶的构象改变,更接近人类4-硫酸酯酶的结构)。一种单克隆抗体选择性地与人4-硫酸酯酶反应,建立了一种适合于评价猫组织中人4-硫酸酯酶替代的测定方法。
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Immunochemical Characterization of Feline and Human N-Acetylgalactosamine 4-Sulfatase

Maroteaux-Lamy syndrome (mucopolysaccharidosis type VI; MPS VI) is a disorder which results from a deficiency in the lysosomal associated enzyme N-acetylgalactosamine 4-sulfatase (4-sulfatase). A feline model of human MPS VI has previously been described and provides a system for the evaluation of enzyme replacement therapy protocols. As a preliminary study to human 4-sulfatase enzyme replacement therapy in feline we have compared the immunochemical properties of human and feline 4-sulfatase. By SDS-PAGE the molecular mass of purified feline and human 4-sulfatase were similar under both reducing and nonreducing conditions. There was, however, a detectable conformation difference between human and feline 4-sulfatase indicating some structural variation. Feline 4-sulfatase reacted weakly with a panel of monoclonal antibodies in an immunobinding assay (interacting with 4-sulfatase in free solution), but the same monoclonal antibodies reacted strongly with feline 4-sulfatase in an immunoquantification assay where the feline 4-sulfatase was bound to a polyclonal antibody (which presumably induces a conformation change in the feline 4-sulfatase to closer approximate the structure of human 4-sulfatase). A monoclonal antibody which selectively reacts with human 4-sulfatase has been used to develop an assay suitable for evaluating human 4-sulfatase enzyme replacement in cat tissues.

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