Anti-HIV Activity of Lung SRCR (gp-340) in Saliva

描述(申请人提供)：虽然从HIV阳性受试者的唾液中获得的HIV RNA序列通常是PCR阳性的，但从口腔样本中培养病毒是极其困难的。再加上流行病学的观察，艾滋病毒很少通过口服传播，现在人们普遍认为唾液中含有使病毒灭活的因素。我们先前证明，大多数人的唾液在体外能够特异性地抑制HIV-1感染。从表现出高水平抗HIV活性的个体的颌下和腮腺唾液样本被用来鉴定和纯化两种高分子量唾液酸糖蛋白，唾液凝集素(SAG)和粘蛋白(MG2)。这两种纯化的糖蛋白在体外都能抑制在唾液中发现的浓度的HIV-1感染。我们随后证明，抑制作用的机制包括唾液蛋白与gp120结合，以及随后gp120从病毒粒子中脱落。 Prakobphol等人(2000)利用蛋白质组学技术证明了SAG与肺清道夫受体蛋白GP-340相同，GP-340是清道夫受体富含半胱氨酸(SRCR)家族的成员。我们获得了该基因的分子克隆，并在哺乳动物细胞中表达了该蛋白，发现重组GP-340在结构和生物学活性上与SAG完全相同。在本提案中，我们将(1)确定gp120上与GP-340相互作用的区域；创建稳定的高水平表达GP-340的转基因细胞，并证明其在所有生物学性质上确实与SAG相同；(2)鉴定GP-340调节env结合和抑制艾滋病毒的离散区域；以及(3)建立GP-340生物活性部分相互作用的机制(S)，以便开发具有抗艾滋病毒活性的新型治疗药物。

DESCRIPTION (provided by applicant): While saliva obtained from HIV positive subjects is often PCR positive for HIV RNA sequences, it is extremely difficult to culture virus from oral samples. Coupled with the epidemiologic observations that HIV is rarely orally transmitted, it is now widely believed that saliva contains factors that inactivate the virus. We previously demonstrated that saliva from most individuals was able to specifically inhibit HIV-1 infection in vitro. Submandibular and parotid saliva samples from individuals demonstrating high levels of anti-HIV activity were used to identify and then purify two high molecular weight sialyated glycoproteins, salivary agglutinin (SAG) and mucin (MG2). Both of these purified glycoproteins inhibited HIV-1 infection in vitro at concentrations found in saliva. We subsequently demonstrated that the mechanism of inhibition involved binding of the salivary proteins to gp120, and subsequent shedding of gp120 from the virion. Prakobphol et al (2000), utilizing proteomic technology, demonstrated that SAG was identical to the lung scavenger receptor protein, gp-340, a member of the scavenger receptor cysteine rich (SRCR) family. We obtained a molecular clone for this gene, expressed the protein in mammalian cells and found that recombinant gp-340 was identical to SAG in terms of its structural and biologic activities. In the present proposal we will (1) Identify regions on gp120 that interact with gp-340; create stable transfected cells that express high levels of gp-340, and demonstrate that it is indeed identical to SAG in all of its biologic properties; (2) Identify discrete regions of gp-340 that mediate env binding and HIV inhibition; and (3) establish the mechanism(s) involved in the interaction of the bioactive portion of gp-340 in order to develop novel therapeutic agents with anti-HIV activity.