Construction and structural characterization of versatile lactosaminoglycan-related compound library for the synthesis of complex glycopeptides and glycosphingolipids.
Construction and structural characterization of versatile lactosaminoglycan-related compound library for the synthesis of complex glycopeptides and glycosphingolipids.
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用于合成复杂糖肽和糖鞘脂的多功能乳糖胺聚糖相关化合物库的构建和结构表征。
DOI：
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发表时间：
2006
影响因子：
3.6
通讯作者：
S. Nishimura
中科院分区：
文献类型：
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作者：
K. Naruchi;T. Hamamoto;Masaki Kurogochi;H. Hinou;H. Shimizu;Takahiko Matsushita;N. Fujitani;H. Kondo;S. Nishimura
我们已经建立了一个简便和有效的协议，用于批量合成各种化合物库相关的乳糖胺聚糖：细胞表面寡糖组成的N-乙酰乳糖胺作为一个重复的二糖单元，基于化学和酶的方法。研究了脑膜炎奈瑟氏菌β 1，3-N-乙酰葡糖胺转移酶（LgtA）的底物特异性和可行性，以合成适合于构建含聚-N-乙酰乳糖胺结构的哺乳动物O-糖肽和鞘糖脂的各种关键中间体。重组LgtA在强碱性条件（pH = 10，甘氨酸-NaOH缓冲液）和20至30 ℃的宽范围的最佳温度下表现出最高的糖基转移酶活性。有趣的是，发现LgtA区分L-丝氨酸和L-苏氨酸，并且作为核心-1 β 1，3-N-乙酰葡糖胺基转移酶和核心-2 β 1，3-N-乙酰葡糖胺基转移酶两者对Fmoc-Ser衍生物起作用，而LgtA在Fmoc-Thr衍生物存在下仅显示核心-2 β 1，3-N-乙酰葡糖胺基转移酶活性。LgtA与人β 1，4-半乳糖基转移酶的组合使用允许从合成糖氨基酸进行受控的糖延伸反应，并得到合成的乳糖胺聚糖，例如十糖衍生物，Galbeta（1 -> 4）GlcNAc beta（1 -> 3）Galbeta（1 -> 4）GlcNAc beta（1 -> 3）Galbeta（1 -> 4）GlcNAc beta（1 -> 3）Galbeta（1 -> 4）GlcNAc beta（1 -> 6）[Galbeta（1 -> 6）] 3）] GalNA α 1-> Fmoc-Ser-OH（6），和十二糖衍生物，Galbeta（1 --> 4）GlcNAc beta（1 --> 3）Galbeta（1 --> 4）GlcNAc beta（1 --> 3）Galbeta（1 --> 4）GlcNAc beta（1 --> 6）[Galbeta（1 --> 4）GlcNAc beta（1 --> 3）Galbeta（1 --> 4）GlcNAc beta（1 --> 3）] GalNA α 1-> Fmoc-Ser-OH（9）。将部分保护的五糖中间体GlcNAc beta（1 -> 3）Galbeta（1 -> 4）GlcNAc beta（1 -> 6）[Galbeta（1 -> 3）] GalNAcalpha 1-> Fmoc-Thr-OH（11）用于MUC 1相关糖肽19（MW = 2610.1）的微波辅助固相合成。研究结果表明，这种糖延伸策略可以用于修饰乳糖基神经酰胺模拟聚合物，以提供方便的前体用于合成各种鞘糖脂。
We have established a facile and efficient protocol for the preparative-scale synthesis of various compound libraries related to lactosaminoglycans: cell surface oligosaccharides composed of N-acetyllactosamine as a repeating disaccharide unit, based on chemical and enzymatic approaches. Substrate specificity and feasibility of a bacterial glycosyltransferase, Neisseria meningitidis beta1,3-N-acetylglucosaminyltransferase (LgtA), were investigated in order to synthesize various key intermediates suited for the construction of mammalian O-glycopeptides and glycosphingolipids containing poly-N-acetyllactosamine structures. Recombinant LgtA exhibited the highest glycosyltransferase activity with strongly basic conditions (pH = 10, glycine-NaOH buffer) and a broad range of optimal temperatures from 20 to 30 degrees C. Interestingly, it was found that LgtA discriminates L-serine and L-threonine and functions both as a core-1 beta1,3-N-acetylglucosaminyltransferase and core-2 beta1,3-N-acetylglucosaminyltransferase toward Fmoc-Ser derivatives, while LgtA showed only core-2 beta1,3-N-acetylglucosaminyltransferase activity in the presence of Fmoc-Thr derivatives. Combined use of LgtA with human beta1,4-galactosyltransferase allowed for controlled sugar extension reactions from synthetic sugar amino acids and gave synthetic lactosaminoglycans, such as a decasaccharide derivative, Galbeta(1 --> 4)GlcNAcbeta(1 --> 3)Galbeta(1 --> 4)GlcNAcbeta(1 --> 3)Galbeta(1 --> 4)GlcNAcbeta(1 --> 3)Galbeta(1 --> 4)GlcNAcbeta(1 --> 6)[Galbeta(1 --> 3)]GalNAcalpha1 --> Fmoc-Ser-OH (6), and a dodecasaccharide derivative, Galbeta(1 --> 4)GlcNAcbeta(1 --> 3)Galbeta(1 --> 4)GlcNAcbeta(1 --> 3)Galbeta(1 --> 4)GlcNAcbeta(1 --> 6)[Galbeta(1 --> 4)GlcNAcbeta(1 --> 3)Galbeta(1 --> 4)GlcNAcbeta(1 --> 3)Galbeta(1 --> 3)]GalNAcalpha1 --> Fmoc-Ser-OH (9). A partially protected pentasaccharide intermediate, GlcNAcbeta(1 --> 3)Galbeta(1 --> 4)GlcNAcbeta(1 --> 6)[Galbeta(1 --> 3)]GalNAcalpha1 --> Fmoc-Thr-OH (11), was applied for the microwave-assisted solid-phase synthesis of a MUC1-related glycopeptide 19 (MW = 2610.1). The findings suggest that this sugar extension strategy can be employed for the modification of lactosyl ceramide mimetic polymers to afford convenient precursors for the synthesis of various glycosphingolipids.