DELINEATION OF GP120 BINDING SITE ON GALCER

我们实验室的实验表明，半乳糖神经酰胺 （GalCer或半乳糖苷），或密切相关的分子发挥作用， 在HIV-1进入细胞系中的重要作用， 人类神经系统 在这些研究中，我们证明了抗体 针对GalCer抑制或减少两种细胞系，U373-MG， 来源于胶质母细胞瘤的SK-N-MC和来源于外周血淋巴细胞的SK-N-MC。 神经母细胞瘤 此外，在伴随的实验中， 重组gp 120、HIV受体结合蛋白和 固定在HPTLC板上的GalCer。 这种结合是饱和的， 部分映射到GalCer分子的极性头部，如预期的，如果 这种相互作用将在体内发生。 由于GalCer是一种重要的 神经系统糖脂，约占干重的14% 是脑白色物质的重要组成部分，是髓鞘形成的关键表面分子 细胞，这些结果在解释一些 HIV感染者的神经系统异常。 延长 根据这些发现，我们将确定gp 120的区域 （一）对非政府组织的影响; 通过单特异性或单克隆抗gp 120抗体的结合，（ii） 竞争gp 120-GalCer与肽的结合，和（iii）产生 gp 120突变体。 我们将探讨gp 120-GalCer结合的可能性， 是由于使用凝集素的碳水化合物-碳水化合物相互作用， 抗碳水化合物的抗体，并通过表达重组gp 120， 具有糖基化缺陷的细胞系。 使用体外结合的发现 测定将与SK-N-MC细胞中的病毒感染相关。 这些结果 能让我们更好地了解这种病毒 蛋白质是艾滋病毒致病性的关键决定因素，也是常见的神经元 系统糖脂。

Experiments from our laboratory have demonstrated that galactosyl ceramide (GalCer or galactocerebroside), or a closely related molecule plays a significant role in the entry of HIV-1 into cell lines derived from the human nervous system. In these studies we demonstrated that antibodies against GalCer inhibited or decreased infection of two cell lines, U373-MG, derived from glioblastoma, and SK-N-MC, derived from a peripheral neuroblastoma. Furthermore, in accompanying experiments there was specific binding between recombinant gp120, the HIV receptor binding protein, and GalCer immobilized on an HPTLC plate. This binding was saturable, and was partially mapped to the polar head of the GalCer molecule, as expected if this interaction were to occur in vivo. Since GalCer is an important nervous system glycolipid, comprising around 14 percent of the dry weight of brain white matter, and is a critical surface molecule in myelinating cells, these results are potentially important in explaining some of the nervous system abnormalities noted in HIV infected individuals. To extend these findings, we will define the region(s) of gp120 responsible for this interaction using several complementary approaches: (i) inhibition of binding by monospecific or monoclonal anti-gp120 antibodies, (ii) competition for gp120-GalCer binding with peptides, and (iii) generation of gp120 mutants. We will explore the possibility that gp120-GalCer binding is due to a carbohydrate-carbohydrate interaction by using lectins, antibodies against carbohydrates and by expressing recombinant gp120 in cell lines with glycosylation defects. The findings using in vitro binding assays will be related to virus infection in SK-N-MC cells. These results will give us a better understanding of the interaction of this viral protein, a key determinant of HIV pathogenicity, and a common nervous system glycolipid.