Analysis of mutational lethality in highly conserved amino acids of the HIV-1 Gag

HIV-1 Gag 高度保守氨基酸的突变致死率分析

• 批准号: 7754896
• 负责人: Yi Liu
• 金额: $ 24.9万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-23 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7754896
• 关键词: AIDS Vaccines; Acute; Address; Affect; Amino Acid Sequence; Amino Acids; Anti-Retroviral Agents; Antigens; Autologous; Biological Assay; Cells; Clone Cells; Consensus; Containment; Databases; Elements; Enrollment; Epitopes; Failure; Financial compensation; Gagging; Genetic; Genetic Recombination; Genome; Growth; HIV; HIV vaccine; HIV-1; HIV-1 vaccine; Heel; Immune; Immune response; Immunology; In Vitro; Individual; Infection; Laboratories; Maps; Mono-S; Mutate; Mutation; Nature; Nucleotides; Peptide Sequence Determination; Plasmids; Population; Proteins; Proteome; Relative (related person); Reporting; Role; Site; Site-Directed Mutagenesis; Staining method; Stains; Structure; Symptoms; Testing; Time; Universities; Vaccine Design; Vaccines; Variant; Vertebral column; Viral; Viral Load result; Viral Physiology; Virus; Virus Diseases; Washington; cohort; cost; design; env Gene Products; fitness; follow-up; gag Gene Products; insight; meetings; novel; novel strategies; pandemic disease; pressure; public health relevance; vaccine development

DESCRIPTION (provided by applicant): One major obstacle for HIV vaccine development is the enormous genetic variability of viral strains between and within individuals. Recently, universal HIV-1 vaccine approaches have been proposed that exclusively target the highly conserved regions of the viral proteome, in which mutations are predicted to severely compromise virus viability. The underlining hypothesis is that a single mutation at a highly conserved amino acid site can be lethal or nearly lethal to the infecting virus and that the occurrences of compensation for these mutations are rare, e.g., multiple and/or fitness-reducing secondary mutations are required for compensation. However, the lethality of mutations at highly conserved amino acid sites has yet to be shown. In addition, we have found evidence indicating that the deleterious effect of mutations at highly conserved sites may be compensated for by mutations at variable sites. We define amino acid sites that are over 98% conserved in HIV-1 group M as highly conserved sites (HCS) and others as variable sites (VS). We propose a pilot exploratory study to test the underlining hypothesis of this vaccine approach. Using site-directed mutagenesis and in vitro growth competition assays, we will examine the lethality of naturally occurring Gag and Env mutations at 20 HCS (one mutation at each HCS) observed in an HIV-1 subtype B infected subject (PIC1362). Our objectives are: in PIC1362, 1) determine the lethality of the observed HCS mutations in the Gag and Env proteins, 2) determine the compensatory effect of the observed VS mutations; and 3) in two other HIV-1 infected subjects (PIC1113 and PIC1693), determine the lethality of Gag HCS mutations observed in PIC1362, to provide initial evidence of the generality of our findings in PIC1362. The results will provide new insights into the critical role of highly conserved amino acid sites in HIV-1 structure and function and help define the proteome constituents for inclusion in a conserved-elements HIV vaccine. PUBLIC HEALTH RELEVANCE: A universal HIV-1 vaccine design exclusively targeting the highly conserved regions of the viral proteome may present a novel way to tackle the enormous diversity in the HIV-1 viral population. This proposal tests the underline hypothesis of this approach towards HIV-1 vaccine development.

描述（由申请人提供）：HIV 疫苗开发的一个主要障碍是个体之间和个体内部病毒株的巨大遗传变异性。最近，人们提出了通用的 HIV-1 疫苗方法，专门针对病毒蛋白质组的高度保守区域，预计该区域的突变会严重损害病毒的活力。强调的假设是，高度保守的氨基酸位点的单一突变可能对感染病毒是致命的或几乎致命的，并且这些突变的补偿的发生是罕见的，例如，需要多个和/或适应度降低的二次突变来补偿。然而，高度保守的氨基酸位点突变的致死性尚未得到证实。此外，我们发现的证据表明，高度保守位点的突变的有害影响可以通过可变位点的突变来补偿。我们将 HIV-1 M 组中保守性超过 98% 的氨基酸位点定义为高度保守位点 (HCS)，将其他位点定义为可变位点 (VS)。我们提出了一项试点探索性研究来测试这种疫苗方法的基本假设。使用定点诱变和体外生长竞争测定，我们将检查在 HIV-1 B 亚型感染受试者 (PIC1362) 中观察到的 20 个 HCS（每个 HCS 一个突变）中自然发生的 Gag 和 Env 突变的致死率。我们的目标是：在PIC1362中，1）确定Gag和Env蛋白中观察到的HCS突变的致死率，2）确定观察到的VS突变的补偿效应； 3) 在另外两名 HIV-1 感染受试者（PIC1113 和 PIC1693）中，确定在 PIC1362 中观察到的 Gag HCS 突变的致死率，为我们在 PIC1362 中的发现的普遍性提供初步证据。这些结果将为高度保守的氨基酸位点在 HIV-1 结构和功能中的关键作用提供新的见解，并有助于定义包含在保守元件 HIV 疫苗中的蛋白质组成分。公共卫生相关性：专门针对病毒蛋白质组高度保守区域的通用 HIV-1 疫苗设计可能会提供一种解决 HIV-1 病毒群体巨大多样性的新方法。该提案测试了这种 HIV-1 疫苗开发方法的基本假设。