Fluharty A L, Meek W E, Katona Z, Tsay K K
University of California, Los Angeles School of Medicine, Lanterman Developmental Center, Pomona 91769.
Biochem Med Metab Biol. 1992 Feb;47(1):86-96. doi: 10.1016/0885-4505(92)90010-v.
Highly purified cerebroside sulfate activator from pig kidneys was characterized by a number of chemical and biological procedures. Methods for chemical modifications were evaluated in an attempt to obtain biologically active derivatives. Iodination, dabsylation, and to a lesser degree reductive methylation provided useful products with good retention of cerebroside sulfate activator activity. Other procedures resulted in largely inactive derivatives or losses in both protein and biological activities. Attempts at renaturation of cerebroside sulfate activator subjected to various denaturing conditions appeared to be successful in many instances, but it was uncertain if the protein structure had actually been disrupted. The binding of cerebroside sulfate by activator was estimated by gel filtration under conditions similar to those of its assay. The formation of a relatively stable 1:1 complex was observed, collaborating results with the human protein. The complex was stable enough to be isolated and shown to be an efficient substrate for arylsulfatase A. The effectiveness of the pig kidney cerebroside sulfate activator for correcting the metabolic defect in activator-deficient human fibroblasts was compared with human materials. The pig kidney protein was taken up more efficiently by the cells and resulted in a better metabolic correction than material from human liver, but was somewhat less effective than a preparation from human urine.
通过多种化学和生物学方法对从猪肾中高度纯化的硫酸脑苷脂激活剂进行了表征。对化学修饰方法进行了评估，试图获得具有生物活性的衍生物。碘化、丹磺酰化以及程度稍低的还原甲基化提供了具有良好硫酸脑苷脂激活剂活性保留率的有用产物。其他方法大多导致衍生物无活性或蛋白质和生物活性均丧失。对处于各种变性条件下的硫酸脑苷脂激活剂进行复性的尝试在许多情况下似乎是成功的，但不确定蛋白质结构是否真的被破坏。在与其测定条件相似的条件下，通过凝胶过滤估计激活剂与硫酸脑苷脂的结合。观察到形成了相对稳定的1:1复合物，这与人源蛋白质的结果一致。该复合物足够稳定，可以分离出来，并被证明是芳基硫酸酯酶A的有效底物。将猪肾硫酸脑苷脂激活剂纠正激活剂缺陷型人成纤维细胞代谢缺陷的有效性与人源材料进行了比较。猪肾蛋白被细胞更有效地摄取，并且比人肝材料产生更好的代谢纠正，但比人尿制剂的效果稍差。
