Kolbinger F, Streiff M B, Katopodis A G
Novartis Pharma AG, Transplantation Preclinical Research, CH 4002 Basel, Switzerland.
J Biol Chem. 1998 Jan 2;273(1):433-40. doi: 10.1074/jbc.273.1.433.
Biochemical evidence suggests that the galactosyltransferase activity synthesizing type 1 carbohydrate chains is separate from the well characterized enzyme that is responsible for the synthesis of type 2 chains. This was recently confirmed by the cloning, from melanoma cells, of an enzyme capable of synthesizing type 1 chains, which was shown to have no homology to other galactosyltransferases. We report here the molecular cloning and functional expression of a second human beta3-galactosyltransferase distinct from the melanoma enzyme. The new beta3-galactosyltransferase has homology to the melanoma enzyme in the putative catalytic domain, but has longer cytoplasmic and stem regions and a carboxyl-terminal extension. Northern blots showed that the new gene is present primarily in brain and heart. When transfected into mammalian cells, this gene directs the synthesis of type 1 chains as determined by a monoclonal antibody specific for sialyl Lewisa. A soluble version of the cloned enzyme was expressed in insect cells and purified. The soluble enzyme readily catalyzes the transfer of galactose to GlcNAc to form Gal(beta1-3)GlcNAc. It also has a minor but distinct transfer activity toward Gal, LacNAc, and lactose, but is inactive toward GalNAc.
生化证据表明，合成1型碳水化合物链的半乳糖基转移酶活性与负责合成2型链的已充分表征的酶是分开的。最近，通过从黑色素瘤细胞中克隆出一种能够合成1型链的酶，证实了这一点，该酶与其他半乳糖基转移酶没有同源性。我们在此报告了另一种不同于黑色素瘤酶的人β3 - 半乳糖基转移酶的分子克隆和功能表达。新的β3 - 半乳糖基转移酶在推定的催化结构域与黑色素瘤酶具有同源性，但具有更长的细胞质和茎区域以及羧基末端延伸。Northern印迹显示新基因主要存在于脑和心脏中。当转染到哺乳动物细胞中时，该基因指导合成1型链，这是通过对唾液酸化路易斯a特异的单克隆抗体确定的。克隆酶的可溶性形式在昆虫细胞中表达并纯化。可溶性酶很容易催化半乳糖转移到GlcNAc上形成Gal(β1 - 3)GlcNAc。它对Gal、LacNAc和乳糖也有轻微但明显的转移活性，但对GalNAc无活性。
