GALACTOSYLTRANSFERASES--STRUCTURE AND REGULATION

糖基转移酶催化细胞和胞外分子合成特定的寡糖结构，在正常和病理状态下的生物识别过程中起着至关重要的作用；一些糖基转移酶还作为膜结合的碳水化合物受体，另一些糖基转移酶在发育调节中起重要作用。然而，关于这些重要酶的结构、相互作用和机制的信息很少。将研究两种半乳糖转移酶，它们分别催化被转移的单糖的构型保持和倒置的反应。然而，共享的序列基序表明，这些酶至少有部分可能是远缘关系的。α-半乳糖基转移酶催化合成半乳糖-α-1，3-半乳糖-β-OR结构；这种酶在人类体内失活，大约1%的内源性抗体是针对其作用产物-α-半乳糖表位的，是异种器官移植的主要障碍。这种酶的机制及其结构基础将通过对衍射率为2.0埃单位的可用晶体进行动力学、突变和协同X射线衍射分析来研究。这些结果将提供有关催化A型和B型血多糖和Forrssman糖脂合成的同源酶的特异性基础的信息。β-4-半乳糖基转移酶-1是人体内六种同源酶之一，催化β-半乳糖在糖蛋白和糖脂中转移到N-乙酰氨基葡萄糖或葡萄糖。α-乳清蛋白是一种与溶菌酶同源的乳腺特异蛋白质，它的结合调节了酶的特异性，使其能够在哺乳期催化乳糖的生物合成(半乳糖转移到葡萄糖)。结构和突变方法将被用来研究通过这种蛋白质-蛋白质相互作用调节的基础。一种新的α-乳清蛋白和溶菌酶的同源物被鉴定为睾丸表达的cDNA.它缺少溶菌酶催化位点，与α-乳清蛋白有一些相似之处，这增加了它可能是一种新的糖基转移酶调节蛋白的可能性。将使用该蛋白的重组形式来研究其活性和结构。

Glycosyltransferases catalyze the synthesis of specific oligosaccharide structures on cells and extracellular molecules that have crucial roles in biological recognition processes in normal and pathological states; some also act as membrane-bound carbohydrate receptors and others are important in the regulation of development. However, there is little information available on the structures, interactions, and mechanisms of these important enzymes. Two galactosyltransferases will be investigated that catalyze reactions in which the configuration of the transferred monosaccharide is retained and inverted, respectively. However, shared sequence motifs suggest that, at least parts of these enzymes may be distantly related. Alpha- Galactosyltransferase catalyze the synthesis of a galactosyl- alpha-1,3-galactosyl-beta-OR structure; this enzyme is inactivated in humans and about 1 percent of endogenous antibodies are directed against the product of its action, the alpha-gal epitope, and represent a major barrier to the xenotransplantation of organs. The mechanism of this enzyme and its structural basis will be investigated by kinetic, mutational and collaborative X-ray diffraction analysis of available crystals that diffract to 2.0 Angstrom units resolution. The results will provide information about the basis of specificity in homologous enzymes that catalyze the synthesis of blood group A and B glycans and Forrssman glycolipid. Beta-4- Galactosyltransferase-1, is one of six homologous enzymes in humans that catalyze the transfer of beta-galactose to N- acetylglucosamine or glucose in glycoproteins and glycolipids. The binding of alpha-lactalbumin, a mammary specific protein that is homologous with lysozyme, modulates the specificity of the enzyme to allow it to catalyze lactose biosynthesis (galactose transfer to glucose) during lactation. Structural and mutational approaches will be used to investigate the basis of regulation through this protein-protein interaction. A new homologue of alpha-lactalbumin and lysozyme has been identified as a testis- expressed cDNA. This lacks the lysozyme catalytic site and has some similarities to alpha-lactalbumin, raising the possibility that it could be a novel glycosyltransferase regulatory protein. The activity and structure will be investigated using a recombinant form of this protein.