Oligosaccharide moieties of human milk glycans that inhibit pathogens

描述（由申请人提供）：现在有强有力的数据支持我们早期的假设，即母乳喂养婴儿肠道疾病发病率的降低部分是由于人乳聚糖的保护。聚糖，包括糖蛋白、糖脂、粘蛋白和糖胺聚糖，含有附着在蛋白质、脂质和其他分子骨架上的复杂低聚糖结构。这些复杂的碳水化合物部分是由乳腺中的许多糖基转移酶合成的；那些与细胞表面糖结合病原体受体同源的聚糖可以抑制病原体结合，从而保护哺乳婴儿。轮状病毒和HIV感染主要通过口服接种引入婴儿，我们发现这些致病性病毒在体外被特定的高分子量人乳聚糖强烈抑制。人乳糖胺聚糖的一个子集、硫酸软骨素和人乳硫酸糖鞘脂部分的一个成分在固相测定和人白细胞中都能强烈抑制HIV。人乳糖蛋白乳酸粘附素（46 kDa）对ma104细胞的轮状病毒感染有很强的抑制作用，这一分子是人乳对轮状病毒体外抑制活性的全部原因。本提案的重点是表征这些活性抗病毒聚糖，使用最新的仪器分析技术，可以提供复杂聚糖部分分子结构的详细了解。研究其分子机制，包括整个分子对多种病原菌的抑制作用。接下来的研究将确定活性分子的最小活性部分，并确定其抑制病原体的机制。这些研究的最终目的是通过克鲁维酵母的基因工程合成活性部分，这种酵母天然存在于人类饮食的许多乳制品中，并且已被证明可以合成其他活性的人乳聚糖。可用于口服、可抑制艾滋病毒或轮状病毒的合成人乳聚糖的可用性可能对这些对公众健康的重要威胁产生重大影响。

DESCRIPTION (provided by applicant): There is now strong data supporting our early hypothesis that the reduced incidence of enteric diseases in breastfeeding infants is due, in part, to protection by human milk glycans. Glycans, including glycoproteins, glycolipids, mucins, and glycosaminoglycans, contain complex oligosaccharide structures attached to proteins, lipids, and other molecular backbones. These complex carbohydrate moieties are synthesized by the many glycosyltransferases in the mammary gland; those glycans with homology to cell surface glycoconjugate pathogen receptors may inhibit pathogen binding, thereby protecting the nursing infant. Rotavirus and HIV infections in infants are introduced primarily through oral inoculation, and we discovered that these pathogenic viruses are strongly inhibited by specific high molecular weight human milk glycans in vitro. A subset of the human milk glycosaminoglycans, a chondroitin sulfate, and a component of the human milk sulfated glycosphingolipid fraction each strongly inhibit HIV in solid phase assays and in human leukocytes. A human milk glycoprotein, lactadherin (46 kDa), strongly inhibits rotavirus infection of MA 104 cells, and this molecule accounts for all of the inhibitory activity of human milk against rotaviruses in vitro. This proposal focuses on characterizing these actively antiviral glycans, using state-of-the-art instrumental analytical techniques that have recently become available and can provide detailed understanding of the molecular structure of complex glycan moieties. The molecular mechanisms will be studied to include the inhibition of multiple strains of the pathogens by the whole molecules. This will be followed by research to identify the smallest active moieties of the active molecules, and confirming their mechanism of pathogen inhibition. These studies are designed to culminate in a plan to synthesize the active moieties through genetic engineering of Kluyveromyces lactis, yeast occurring naturally in many dairy products of the human diet, and which has proved amenable to synthesis of other active human milk glycans. The availability of synthetic human milk glycans suitable for oral consumption that inhibit HIV or rotavirus could have large impact on these important threats to public health.