Joziasse D H, Schiphorst W E, Van den Eijnden D H, Van Kuik J A, Van Halbeek H, Vliegenthart J F
J Biol Chem. 1987 Feb 15;262(5):2025-33.
Using 500-MHz 1H NMR spectroscopy we have investigated the branch specificity that bovine colostrum CMP-NeuAc:Gal beta 1----4GlcNAc-R alpha 2----6-sialyltransferase shows in its sialylation of bi-, tri-, and tetraantennary glycopeptides and oligosaccharides of the N-acetyllactosamine type. The enzyme appears to highly prefer the galactose residue at the Gal beta 1----4GlcNAc beta 1----2Man alpha 1----3 branch for attachment of the 1st mol of sialic acid in all the acceptors tested. The 2nd mol of sialic acid becomes linked mainly to the Gal beta 1----4GlcNAc beta 1----2Man alpha 1----6 branch in bi- and triantennary substrates, but this reaction invariably proceeds at a much lower rate. Under the conditions employed, the Gal beta 1----4GlcNAc beta 1----6Man alpha 1----6 branch is extremely resistant to alpha 2----6-sialylation. A higher degree of branching of the acceptors leads to a decrease in the rate of sialylation. In particular, the presence of the Gal beta 1----4GlcNAc beta 1----6Man alpha 1----6 branch strongly inhibits the rate of transfer of both the 1st and the 2nd mol of sialic acid. In addition, it directs the incorporation of the 2nd mol into tetraantennary structures toward the Gal beta 1----4GlcNAc beta 1----4Man alpha 1----3 branch. In contrast, the presence of the Gal beta 1----4GlcNAc beta 1----4Man alpha 1----3 branch has only minor effects on the rates of sialylation and, consequently, on the branch preference of sialic acid attachment. Results obtained with partial structures of tetraantennary acceptors indicate that the Man beta 1----4GlcNAc part of the core is essential for the expression of branch specificity of the sialyltransferase. The sialylation patterns observed in vivo in glycoproteins of different origin are consistent with the in vitro preference of alpha 2----6-sialyltransferase for the Gal beta 1----4GlcNAc beta 1----2Man alpha 1----3 branch. Our findings suggest that the terminal structures of branched glycans of the N-acetyllactosamine type are the result of the complementary branch specificity of the various glycosyltransferases that are specific for the acceptor sequence Gal beta 1----4GlcNAc-R.
我们使用500兆赫的1H核磁共振光谱，研究了牛初乳CMP-唾液酸：β-1,4-半乳糖基-N-乙酰葡糖胺-R α2,6-唾液酸转移酶在对N-乙酰乳糖胺型二天线、三天线和四天线糖肽及寡糖进行唾液酸化时所表现出的分支特异性。在所有测试的受体中，该酶似乎高度优先选择β-1,4-半乳糖基-β-1,2-甘露糖基-α-1,3分支上的半乳糖残基来连接第一摩尔唾液酸。第二摩尔唾液酸主要连接到二天线和三天线底物的β-1,4-半乳糖基-β-1,2-甘露糖基-α-1,6分支上，但此反应的速率始终要低得多。在所采用的条件下，β-1,4-半乳糖基-β-1,6-甘露糖基-α-1,6分支对α2,6-唾液酸化具有极强的抗性。受体分支程度的提高会导致唾液酸化速率降低。特别是，β-1,4-半乳糖基-β-1,6-甘露糖基-α-1,6分支的存在强烈抑制了第一摩尔和第二摩尔唾液酸的转移速率。此外，它将第二摩尔唾液酸掺入四天线结构的过程导向β-1,4-半乳糖基-β-1,4-甘露糖基-α-1,3分支。相反，β-1,4-半乳糖基-β-1,4-甘露糖基-α-1,3分支的存在对唾液酸化速率，进而对唾液酸连接的分支偏好只有轻微影响。用四天线受体的部分结构获得的结果表明，核心的甘露糖-β-键-1,4-N-乙酰葡糖胺部分对于唾液酸转移酶分支特异性的表达至关重要。在体内不同来源糖蛋白中观察到的唾液酸化模式与体外α2,6-唾液酸转移酶对β-1,4-半乳糖基-β-1,2-甘露糖基-α-1,3分支的偏好一致。我们的研究结果表明，N-乙酰乳糖胺型分支聚糖的末端结构是各种对受体序列β-1,4-半乳糖基-N-乙酰葡糖胺-R具有特异性的糖基转移酶互补分支特异性的结果。
