Waheed A, Van Etten R L
Arch Biochem Biophys. 1985 Nov 15;243(1):274-83. doi: 10.1016/0003-9861(85)90796-9.
The biosynthesis of lysosomal acid phosphatase was studied in a normal human embryonic lung cell line, WI-38. Cells were labeled with radioactive leucine under a variety of conditions, the enzyme was immunoprecipitated using a monospecific antiserum raised against human liver lysosomal acid phosphatase, and the products were separated by electrophoresis and were visualized by fluorography. Lysosomal acid phosphatase constitutes 60% of the total tartrate-inhibitable acid phosphatase in WI-38. It is initially synthesized as a high-molecular-weight precursor polypeptide of 69 kDa. The precursor polypeptide is rapidly glycosylated and processed to a mature enzyme of 53-45 kDa via intermediates of 65 and 60 kDa in WI-38 cells. The 69-kDa precursor polypeptide is also converted to larger precursor polypeptides of 74 and 80 kDa. The multiplicity of precursor polypeptides is due at least in part to differences in the glycosylation and phosphorylation of the polypeptides. Sensitivity of phosphorylated oligosaccharide chains from precursor, mature and small polypeptides to endo-beta-hexosaminidase H-catalyzed cleavage suggests the presence of high-mannose phosphorylated oligosaccharide chains similar to those present on many other lysosomal enzymes. The effects of tunicamycin and ammonium chloride were also studied. In contrast to the effect of ammonium chloride on arylsulfatase A secretion, the lysosomal acid phosphatase in WI-38 cells was not secreted in the presence of NH4Cl. This is consistent with the existence of an alternate route for the transfer of lysosomal acid phosphatase into lysosomes. This alternate route may be the reason that I-cell fibroblasts contain a normal level of lysosomal acid phosphatase.
在正常人类胚胎肺细胞系WI-38中研究了溶酶体酸性磷酸酶的生物合成。在多种条件下用放射性亮氨酸标记细胞，使用针对人肝脏溶酶体酸性磷酸酶产生的单特异性抗血清对该酶进行免疫沉淀，产物通过电泳分离并通过荧光自显影进行可视化。溶酶体酸性磷酸酶占WI-38中总酒石酸抑制性酸性磷酸酶的60%。它最初被合成为一种69 kDa的高分子量前体多肽。在WI-38细胞中，前体多肽迅速糖基化并通过65 kDa和60 kDa的中间体加工成53 - 45 kDa的成熟酶。69 kDa的前体多肽也会转化为74 kDa和80 kDa的更大前体多肽。前体多肽的多样性至少部分归因于多肽糖基化和磷酸化的差异。前体、成熟和小多肽的磷酸化寡糖链对内切β - 己糖胺酶H催化裂解的敏感性表明存在与许多其他溶酶体酶上存在的类似的高甘露糖磷酸化寡糖链。还研究了衣霉素和氯化铵的作用。与氯化铵对芳基硫酸酯酶A分泌的影响相反，在存在NH4Cl的情况下，WI-38细胞中的溶酶体酸性磷酸酶不会分泌。这与溶酶体酸性磷酸酶转移到溶酶体中的另一条途径的存在是一致的。这条替代途径可能是I型细胞成纤维细胞含有正常水平溶酶体酸性磷酸酶的原因。
