Seko A, Kitajima K, Iwamatsu T, Inoue Y, Inoue S
Department of Biochemistry, Sasaki Institute, Kanda-Surugadai, Chiyoda-ku, Tokyo 101-0062, Japan.
Glycobiology. 1999 Sep;9(9):887-95. doi: 10.1093/glycob/9.9.887.
Two different types of peptide:N-glycanase (PNGase) were identified in developing embryos of medaka fish ( Oryzias latipes ). Because the optimum pH values for their activities were acidic and neutral, they were designated as acid PNGase M and neutral PNGase M, respectively. The acid PNGase M corresponded to the enzyme that had been partially purified from medaka embryos (Seko,A., Kitajima,K., Inoue,Y. and Inoue,S. (1991) J. Biol. Chem., 266, 22110-22114). The apparent molecular weight of this enzyme was 150 K, and the optimal pH was 3.5-4.0, and the K m for L-hyosophorin was 44 microM. L-Hyosophorin is a cortical alveolus-derived glycononapeptide with a large N-linked glycan chain present in the perivitelline space of the developing embryo. Acid PNGase M was competitively inhibited by a free de-N-glycosylated nonapeptide derived from L-hyosophorin. This enzyme was expressed in ovaries and embryos at all developmental stages after gastrulation, but activity was not detected in embryos at developmental stages between fertilization and gastrula. Several independent lines of evidence suggested that acid PNGase M may be responsible for the unusual accumulation of free N-glycans derived from yolk glycoproteins (Iwasaki,M., Seko,A., Kitajima,K., Inoue,Y. and Inoue,S. (1992) J. Biol. Chem., 267, 24287-24296). In contrast, the neutral PNGase M was expressed in blastoderms from the 4-8 cell stage and in cells up to early gastrula. The general significance of these findings is that they show a developmental stage-dependent expression of the two PNGase activities, and that expression of the neutral PNGase M activity occurs concomitantly with the de-N-glycosylation of L-hyosophorin. These data thus support our conclusion that the neutral PNGase M is responsible for the developmental-stage-related de-N-glycosylation of the L-hyosophorin.
在青鳉鱼（日本青鳉）发育的胚胎中鉴定出了两种不同类型的肽：N-聚糖酶（PNGase）。由于它们活性的最适pH值分别为酸性和中性，因此分别被命名为酸性PNGase M和中性PNGase M。酸性PNGase M与之前从青鳉胚胎中部分纯化得到的酶相对应（Seko，A.，北岛，K.，井上，Y.和井上，S.（1991年）《生物化学杂志》，266，22110 - 22114）。这种酶的表观分子量为150 K，最适pH为3.5 - 4.0，对L-透明带蛋白的K m为44 microM。L-透明带蛋白是一种源自皮质颗粒的糖基九肽，在发育中胚胎的卵周隙中存在一条大的N-连接聚糖链。酸性PNGase M受到源自L-透明带蛋白的游离去N-糖基化九肽的竞争性抑制。这种酶在原肠胚形成后的所有发育阶段的卵巢和胚胎中都有表达，但在受精和原肠胚形成之间的发育阶段的胚胎中未检测到活性。几条独立的证据表明，酸性PNGase M可能是导致源自卵黄糖蛋白的游离N-聚糖异常积累的原因（岩崎，M.，Seko，A.，北岛，K.，井上，Y.和井上，S.（1992年）《生物化学杂志》，267，24287 - 24296）。相比之下，中性PNGase M在4 - 8细胞期的胚盘以及直至早期原肠胚的细胞中表达。这些发现的普遍意义在于，它们显示了两种PNGase活性的发育阶段依赖性表达，并且中性PNGase M活性的表达与L-透明带蛋白的去N-糖基化同时发生。因此，这些数据支持了我们的结论，即中性PNGase M负责L-透明带蛋白与发育阶段相关的去N-糖基化。
