Nauseef W M
Blood. 1986 Apr;67(4):865-72.
The biosynthesis and processing of myeloperoxidase (MPO), a cationic enzyme present in the azurophilic granules of human polymorphonuclear leukocytes (PMNs), were studied in the human promyelocytic leukemia cell line, HL-60. HL-60 cells produce large quantities of enzymatically active MPO that has the same electrophoretic behavior as MPO isolated from normal PMNs. Mature MPO is a glycoprotein of approximately 150,000 molecular weight (mol wt) composed of two heavy-light protomers (alpha 2 beta 2) with subunits of 59,000 and 13,500 mol wt, respectively, under reducing conditions. The primary translation product of MPO messenger RNA (mRNA) isolated from HL-60 cells was a single polypeptide of mol wt 80,000. In HL-60 cells labeled with [35S]-methionine, the labeled MPO isolated by immunoprecipitation had a mol wt of 89,000. Treatment of this 89-kilodalton (kDa) species with endoglycosidase H produced a 79-kDa peptide, suggesting that the 89-kDa protein contained high-mannose side chains. The 89-kDa species had no detectable peroxidase activity. During chase experiments some of the 89-kDa peptide was processed to smaller species of mol wt 39,000, 59,000, and 13,500, although a fraction of the 89-kDa peptide remained unprocessed after a chase of 100 hours. In addition, a small amount of the 89-kDa peptide appeared in the medium without any of the processed smaller peptides. These studies suggest that the primary translation product in MPO biosynthesis is an 80-kDa peptide that undergoes cotranslational cleavage of the signal peptide and glycosylation to produce an 89-kDa pro-MPO, that pro-MPO is a single polypeptide containing the alpha and beta subunits of MPO and contains endoglycosidase H-susceptible high-mannose side chains, and that posttranslational modification of pro-MPO results in targeting to the lysosome and proteolytic maturation of pro-MPO to active enzyme. In light of the previous observation that MPO-deficient and normal PMNs contain an 89-kDa protein immunochemically related to MPO, these studies on MPO biosynthesis indirectly support the hypothesis that defective posttranslation processing by pro-MPO may underlie hereditary MPO deficiency.
在人早幼粒细胞白血病细胞系HL - 60中研究了髓过氧化物酶（MPO）的生物合成与加工过程，MPO是一种存在于人类多形核白细胞（PMN）嗜天青颗粒中的阳离子酶。HL - 60细胞产生大量具有酶活性的MPO，其电泳行为与从正常PMN中分离出的MPO相同。成熟的MPO是一种分子量约为150,000的糖蛋白，在还原条件下由两个重 - 轻亚基（α2β2）组成，亚基分子量分别为59,000和13,500。从HL - 60细胞中分离出的MPO信使核糖核酸（mRNA）的初级翻译产物是一个分子量为80,000的单一多肽。在用[35S] - 甲硫氨酸标记的HL - 60细胞中，通过免疫沉淀分离出的标记MPO分子量为89,000。用内切糖苷酶H处理这种89千道尔顿（kDa）的蛋白产生了一个79 kDa的肽段，这表明89 kDa的蛋白含有高甘露糖侧链。89 kDa的蛋白没有可检测到的过氧化物酶活性。在追踪实验中，一些89 kDa的肽段被加工成分子量为39,000、59,000和13,500的较小肽段，尽管在追踪100小时后仍有一部分89 kDa的肽段未被加工。此外，少量89 kDa的肽段出现在培养基中，而没有任何加工后的较小肽段。这些研究表明，MPO生物合成中的初级翻译产物是一个80 kDa的肽段，它经过信号肽的共翻译切割和糖基化作用产生一个89 kDa的MPO前体，该前体是一个包含MPO的α和β亚基的单一多肽，并且含有对内切糖苷酶H敏感的高甘露糖侧链，并且MPO前体的翻译后修饰导致其靶向溶酶体并将MPO前体蛋白水解成熟为活性酶。鉴于之前观察到MPO缺陷型和正常PMN中含有一种与MPO免疫化学相关的89 kDa蛋白，这些关于MPO生物合成的研究间接支持了这样一种假说，即MPO前体的翻译后加工缺陷可能是遗传性MPO缺乏的基础。
