Human lung adenocarcinoma α1,3/4-L-fucosyltransferase displays two molecular forms, high substrate affinity for clustered sialyl LacNAc type 1 units as well as mucin core 2 sialyl LacNAc type 2 unit and novel α1,2-L-fucosylating activity
Human lung adenocarcinoma α1,3/4-L-fucosyltransferase displays two molecular forms, high substrate affinity for clustered sialyl LacNAc type 1 units as well as mucin core 2 sialyl LacNAc type 2 unit and novel α1,2-L-fucosylating activity
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DOI：
10.1093/glycob/11.5.353
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发表时间：
2001-05-01
期刊：
影响因子：
4.3
通讯作者：
Matta, KL
中科院分区：
文献类型：
--
作者：
Chandrasekaran, EV;Chawda, R;Matta, KL
人肺肿瘤α1，3/4-L岩藻糖基转移酶(FT)经亲和层析纯化，纯化倍数为2000倍，回收率29%。在Sephacryl S-100HR色谱柱上进一步分离得到两种分子形式(FTA，较大尺寸，具有15%的α1，4-FT活性；FTB，主要形式，活性为85%)。在天然聚丙烯酰胺凝胶电泳法(PAGE)上也发现了这两种活性的电泳率的差异。这两种形式都缺乏典型的α1，2-岩藻糖基化活性，但与新的将Lewis a决定簇转化为Lewis b的α1，2-岩藻糖化能力有关。根据对2-O-Mecal Beta1，3GlcNAcβ-O-Bn的活性百分比，肉汤形式对各种受体表现出相同程度的活性，包括硫酸、唾液酸化或甲基化的LacNAc类型1或2以及粘蛋白核心2受体。然而，FTA和FTB对粘蛋白核心2，3‘-唾液酸基LacNAc的作用能力不同(活性分别为24.2%和40.8%，而2-O-Mealβ1，3GlcNAcβ-O-Bn的活性分别为24.2%和40.8%)。未被取代的LacNAc 1型受体的活性是相应的LacNAc 2型受体的15-20倍。粘蛋白核心2受体中的β-1，4-半乳糖基(甲基、硫酸盐或唾液酸基)上的3-O-取代使这些受体的效率提高了五到八倍。FTA和FTB的最有效受体是3-O-磺基半乳糖β-1，3GlcNAc-β-O-Al(K-m分别为100和47um)。2-O-甲基-1，3GlcNAc-β-O-Bn和3-O-唾液酸半乳糖-1，3GlcNAc-β-O-Bn的K-m(Mm)分别为0.40和2.5(FTA)和0.16和0.67(FTB)。在要求分析时，受体效率降至接近50%(asialo ancrod的K-m：167微米)。唾液酸糖蛋白，如癌胚抗原、胎球蛋白和牛α-酸性糖蛋白是比去唾液酸胎蛋白更好的受体，相反，以NeuAcα-2，3Gal-β1，4GlcNAc-β为主的胎球蛋白三天线糖肽的活性仅为去唾液酸糖多肽的55%(K-m分别为1.43和0.63 mM)。因此，人肺肿瘤α1，3/4-L-FT有可能在N-糖链中产生簇状唾液酸路易斯a和刘易斯b决定子，在粘蛋白核心2结构中产生唾液酸路易斯x决定子。
Human lung tumor alpha1,3/4-L-fucosyltransferase (FT) was purified (2000-fold, 29% recovery) from 290 g of tissue by including a chromatography step on Affinity Gel-GDP. Two molecular forms (FTA, larger size carrying 15% alpha1,4-FT activity; FTB, the major form with 85% activity) were separated by further fractionation on a Sephacryl S-100 HR column. A difference in the electrophoretic mobilities of these two activities was also found on native polyacrylamide gel electrophoresis (PAGE). Both forms were devoid of typical alpha1,2-fucosylating activity but were associated with the novel alpha1,2-fucosylating ability of converting the Lewis a determinant to Lewis b. Based on percentage activity toward 2-O-MeCal beta1,3GlcNAc beta -O-Bn, broth forms exhibited the same extent of activity toward various accepters, which included sulfated, sialylated, or methylated LacNAc type 1 or type 2 as well as mucin core 2 accepters. However, FTA and FTB exhibited a difference in their ability to act on mucin core 2 3'-sialyl LacNAc (activities 24.2% and 40.8%, respectively, as compared to 2-O-MeGal beta1,3GlcNAc beta -O-Bn). The unsubstituted LacNAc type 1 accepters were 15-20 times as active as the corresponding LacNAc type 2 accepters. The 3-O-substitution on the beta1,4-linked Gal (methyl, sulfate, or sialyl) in mucin core 2 accepters increased the efficiency of these accepters five- to eightfold. The most efficient acceptor for FTA and FTB was 3-O-sulfoGal beta1,3GlcNAc beta -O-Al (K-m 100 and 47 muM, respectively). The K-m (mM) values for 2-O-methyl Gal beta1,3GlcNAc beta -O-Bn and 3-O-sialyl Gal beta1,3GlcNAc beta -O-Bn were 0.40 and 2.5 (FTA) and 0.16 and 0.67 (FTB), respectively.The 35-kDa glycoprotein ancrod (from Malayan pit viper venom) containing 36% complex N-glycans with the antennae NeuAc alpha2,3Gal beta1,3GlcNAc beta- acted as the best macromolecular acceptor substrate (K-m: 45 muM), as examined with FTB. On desialylation the acceptor efficiency dropped to similar to 50% (K-m for asialo ancrod: 167 muM). Sialylglyco-proteins, such as carcinoembryonic antigen, fetuin, and bovine alpha1-acid glycoprotein, were better accepters than asialo fetuin, On the contrary, fetuin triantennary glycopeptide containing predominantly NeuAc alpha2,3Gal beta1,4GlcNAc beta- was only 55 % active as compared to the asialo glycopeptide (K-m: 1.43 and 0.63 mM, respectively). Thus, the human lung tumor alpha1,3/4-L-FT has the potential to generate clustered sialyl Lewis a and Lewis b determinants in N-glycans and sialyl Lewis x determinant in mucin core 2 structures.