Waheed A, Hasilik A, Cantz M, von Figura K
Hoppe Seylers Z Physiol Chem. 1982 Feb;363(2):169-78. doi: 10.1515/bchm2.1982.363.1.169.
N-Acetylglucosamine-1-phosphotransferase activity was assayed in human skin fibroblasts using [beta-32P]UDP-N-acetylglucosamine as donor and dephosphorylated beta-N-acetyl-D-hexosaminidase as acceptor. An optimal transfer rate of N-acetylglucosamine 1-phosphate required CDP-choline and ADP in order to inhibit the breakdown of [beta-32P]UDP-N-acetylglucosamine and a combination of leupeptin and iodoacetamide to protect the transferase. The transferase required Mg2 or Mn2. Using doubly labelled UDP-N-acetylglucosamine, simultaneous transfer of N-acetyl-[6-3H]glucosamine and [32P]phosphate to endogenous acceptors was demonstrated. Membranes prepared from fibroblasts from patients with mucolipidosis III were defective in transfer of N-acetylglucosamine 1-phosphate. A residual transferase activity of less than 10% of controls was detectable in fibroblast membranes of eight patients with mucolipidosis III. In membranes from fibroblasts from patients with mucolipidosis II,N-acetylglucosamine-1-phosphotransferase activity was not detectable. Our results indicate that the primary defect in mucolipidoses II and III is a deficiency in N-acetylglucosamine-1-phosphotransferase, the residual activity being higher in mucolipidosis III than in mucolipidosis II.
以[β-32P]UDP-N-乙酰葡糖胺作为供体，去磷酸化的β-N-乙酰-D-己糖胺酶作为受体，在人皮肤成纤维细胞中检测N-乙酰葡糖胺-1-磷酸转移酶活性。N-乙酰葡糖胺1-磷酸的最佳转移速率需要CDP-胆碱和ADP来抑制[β-32P]UDP-N-乙酰葡糖胺的分解，还需要亮抑酶肽和碘乙酰胺的组合来保护转移酶。转移酶需要Mg2或Mn2。使用双标记的UDP-N-乙酰葡糖胺，证明了N-乙酰-[6-3H]葡糖胺和[32P]磷酸同时转移到内源性受体上。从黏脂贮积症III型患者的成纤维细胞制备的膜在N-乙酰葡糖胺1-磷酸的转移方面存在缺陷。在8例黏脂贮积症III型患者的成纤维细胞膜中可检测到残留转移酶活性，其活性低于对照的10%。在黏脂贮积症II型患者的成纤维细胞的膜中，未检测到N-乙酰葡糖胺-1-磷酸转移酶活性。我们的结果表明，黏脂贮积症II型和III型的主要缺陷是N-乙酰葡糖胺-1-磷酸转移酶缺乏，黏脂贮积症III型的残留活性高于黏脂贮积症II型。
