MOLECULAR MECHANISM OF VIRUS NEUTRALIZATION

长期目标是研究病毒的结构 糖蛋白及其参与附着、渗透和 以确定它们如何与抗体分子相互作用， 中和病毒的传染性 尽管病毒的中和作用 经过多年的实验研究，人们对这种分子结构知之甚少。 这一重要的自然宿主对病毒感染的反应的基础。 了解这一点是了解传染性如何的先决条件 病毒-抗体复合物在体内产生，以及它们如何促进 许多病毒感染的疾病过程。 拉克罗斯，一种医学上重要的病毒，被选为研究的模型 抗体中和的机制。 它有两个糖蛋白， 但只有G1参与抗体的合成 中和 将通过肽图谱对G1进行表征，以确定 糖基化位点、二硫键等。单克隆抗体具有 已制备并用于映射八个非重叠抗原位点的G1。 这些抗原位点的确切位置将通过使用 免疫沉淀G1的CNBr肽片段的单特异性抗体， 对每个肽的氨基末端进行测序，并将其与 G1的推导氨基酸序列如他人所提供。 是 提出可能有一种主要的中和机制， 在噬斑测定中中和和测量的病毒量可以是 每个病毒-抗体复合物被抑制的概率之和 在某种程度上与细胞相互作用。 为了确定这一点， 了解拉克罗斯病毒的传染性。 为此将 在存在或不存在以下物质的情况下使用结合渗透和融合测定 促溶酶体剂如氯喹。 病毒与每一个 将在这些试验中检查单克隆抗体，以确定 抗体必须结合在G1上以中和病毒，并确定 在增殖循环中的步骤中，病毒复制被阻断。 的 在转录时或转录前中和的可能性也将 接受检查。 有人提出，在结构的构象变化， G1的表达是由抗体结合诱导的，这是病毒感染所必需的。 中和 激光拉曼光谱学将用于尝试 确定抗体结合或抗体对协同结合 诱导G1的构象变化，通过病毒 膜的

The long-term objectives are to investigate the structure of virus glycoproteins and their involvement in attachment, penetration, and uncoating and to determine how they interact with antibody molecules to neutralize virus infectivity. Altough neutralization of virus has been studied emperically for many years, little is known about the molecular basis for this important natural host response to virus infection. Knowledge of this is a prerequisite to the understanding of how infectious virus-antibody complexes arise in vivo and how they contribute to the disease process of many viral infections. La Crosse, a medically important virus, has been chosen as a model to study the mechanism of antibody neutralization. It has two glycoproteins in its envelope, but only one of them, G1, is involved in antibody neutralization. G1 will be characterized by peptide mapping to determine sites of glycosylation, disulfide bonding, etc. Monoclonal antibodies have been prepared and used to map eight non-overlapping antigenic sites on G1. The exact location of these antigenic sites will be determined by using the monospecific antibodies to immunoprecipitate CNBr peptide fragments of G1, sequencing the amino terminus of each peptide, and correlating this with the deduced amino acid sequence of G1 as provided by others. It is proposed that there may be one main mechanism of neutralization but that the amount of virus neutralized and measured in plaque assays may be the sum of the probabilities that each virus-antibody complex will be inhibited from interacting with the cell at some level. To determine this, more must be known about the infectious entry of La Crosse virus. This will be done using binding penetration, and fusion assays in the presence or absence of lysosomotropic agents such as chloroquine. Virus reacted with each monoclonal antibody will be examined in these assays to determine where antibody must bind on G1 to neutralize virus and to determine at what step(s) in the multiplication cycle the virus replication is blocked. The possibility of neutralization at or just prior to transcription will also be examined. It is proposed that a conformational change in the structure of G1 is induced by antibody binding and that this is necessary for virus neutralization. Laser Raman spectroscopy will be used in attempts to determine if binding of antibody or synergistic binding of antibody pairs induces a conformational changes in G1 that is transmitted across the viral membrane.