Elucidating the Chemistry and Biology of Nucleic Acid Cytidine Deaminases in HIV

阐明 HIV 核酸胞苷脱氨酶的化学和生物学

基本信息

批准号：
8604126
负责人：
Rahul Manu Kohli
金额：
$ 13.2万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-02-15 至 2015-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8604126
关键词：
Acquired Immunodeficiency Syndrome Address Antibodies Antibody Affinity Antibody Diversity B-Cell Lymphomas B-Lymphocytes Binding Biochemical Biological Biological Process Biology Catalysis Chemicals Chemistry Chromosomal translocation Chronic Communicable Diseases Complex Coupled Cytidine Deaminase Cytosine DNA DNA Binding DNA biosynthesis Deaminase Deamination Doctor of Philosophy Enzyme Activation Enzymes Epitopes Evolution Family Family member Generation of Antibody Diversity Generations Genes Genome Goals HIV HIV Genome Immune response Immune system Immunoglobulin Class Switching Immunoglobulin Genes Immunoglobulin Somatic Hypermutation Immunoglobulin Switch Recombination Immunologics Immunology In Vitro Infection Kinetics Lead Lesion Link Malignant Neoplasms Mediating Mentors Methodology Methods Molecular Mutation Nature Non-Hodgkin&apos s Lymphoma Nucleic Acid Binding Nucleic Acids Oligonucleotides Oncogenic Outcome Patients Physicians Physiological Play Polynucleotides Proteins Regulation Research Research Personnel Role Site-Directed Mutagenesis Specificity Structure Testing Training Uracil Variant Vertebral column Viral Work activation-induced cytidine deaminase analog base catalyst enzyme mechanism experience human CEM15 protein immune activation in vivo inhibitor/antagonist insight interest member mutant neoplastic novel nucleobase nucleoside analog nucleotide analog overexpression pathogen preference research study response skills vif Gene Products virology

项目摘要

The primary investigator is an MD/PhD trained infectious diseases physician with an interest in understanding enzymes that generate diversity in host-pathogen interactions. In the proposed work, the PI aims to bring his prior experience in enzyme mechanisms and develop new training through virologic experiments and immunologic studies. A remarkable group of enzymes, the polynucleotide cytidine deaminases of the AID/APOBEC family, play both constructive and destructive roles in struggle against HIV. On one hand, deamination by the family member APOBEC3G interferes with the integrity of the pathogen genome. In turn, HIV has evolved the lentiviral protein Vif as an evasive means to counteract human APOBEC3G. Infection with HIV is also associated with immune activation, which can result in increased expression of a B-cell specific deaminase family member, AID. AID physiologically serves as the chief catalyst governing antibody diversity through the introduction of targeted uracil lesions in antibody variable genes or switch regions which ultimately result in higher affinity antibodies of altered isotype. Aberrant regulation and expression of AID has increasingly been associated with Non-Hodgkins lymphoma, the leading AIDS-defining malignancy in HIV infected patients. Despite the importance of these cytidine deaminases, little is know about the nature of their interaction with their nucleic acid targets. This proposal addresses the hypothesis that the molecular interactions that lead to catalysis and binding of nucleic acids are critical determinants of their proper physiologic function. The studies aim to decipher and perturb these molecular interactions. Initially, structure-based hypotheses will be used to localize the protein determinants of sequence preference and resolve the mode of binding to the nucleic acid backbone. By utilizing novel loop graft mutant enzymes with altered sequence preference, the impact of perturbed sequence specificity on retroviral restriction (APOBEC3G) or antibody diversity and chromosomal translocations (AID) will be explored. To understand catalysis by AID/APOBEC enzymes, nucleoside analogs will be introduced into oligonucleotides via chemical or chemoenzymatic methods and are used to characterize the kinetics of deamination and the inhibition of pro-oncogenic AID activity. Taken together, a full characterization of the AID/APOBEC-nucleic acid complex ¿ binding and catalysis ¿ will provide a molecular basis for the action of this important enzyme family in vitro and in vivo. Through mentored training, the PI will develop the broad based research skills necessary to examine biological and biochemical aspects of diversity generation in host-pathogen interactions upon an ultimate transition to independence.

主要研究者是一位受过医学博士/博士学位训练的传染病医生，对以下领域感兴趣了解在宿主-病原体相互作用中产生多样性的酶在拟议的工作中，PI。旨在利用他之前在酶机制方面的经验，并通过病毒学开展新的培训实验和免疫学研究。 AID/APOBEC 家族的多核苷酸胞苷脱氨酶是一组非凡的酶，一方面，家庭的脱氨基作用在抗击艾滋病毒的斗争中发挥着建设性和破坏性的作用。 APOBEC3G 成员干扰了病原体基因组的完整性，反过来，HIV 又进化出了慢病毒蛋白Vif也可作为对抗人类APOBEC3G感染的逃避手段。与免疫激活相关，可导致 B 细胞特异性脱氨酶表达增加 AID 家族成员在生理上是通过调节抗体多样性的主要催化剂。在抗体可变基因或开关区域中引入靶向尿嘧啶损伤，最终导致 AID 的异常调节和表达已日益受到关注。与非霍奇金淋巴瘤有关，非霍奇金淋巴瘤是艾滋病毒感染者中主要的艾滋病定义恶性肿瘤。尽管这些胞苷脱氨酶很重要，但人们对它们与胞苷脱氨酶相互作用的性质知之甚少。该提案提出了导致分子相互作用的假设。核酸的催化和结合是其正常生理功能的关键决定因素。最初，将使用基于结构的假设来破译和扰乱这些分子相互作用。定位序列偏好的蛋白质决定因素并解析与核酸的结合模式通过利用具有序列偏好的新型环移植突变酶，影响逆转录病毒限制 (APOBEC3G) 或抗体多样性和染色体的序列特异性受到干扰将探索易位 (AID) 以了解 AID/APOBEC 酶、核苷类似物的催化作用。将通过化学或化学酶方法引入寡核苷酸并用于表征脱氨动力学和促癌 AID 活性的抑制结合在一起，形成了完整的结果。 AID/APOBEC-核酸复合物的表征 ¿结合和催化 ¿将提供分子通过指导培训，PI 将成为这一重要酶家族在体外和体内作用的基础。培养检查多样性的生物和生化方面所需的广泛研究技能最终过渡到独立时宿主与病原体相互作用的产生。

项目成果

期刊论文数量（9）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Specificity determinants for autoproteolysis of LexA, a key regulator of bacterial SOS mutagenesis.

LexA 自身蛋白水解的特异性决定因素，LexA 是细菌 SOS 诱变的关键调节因子。

DOI：
发表时间：
2014-05-20
期刊：
影响因子：
2.9
作者：
Mo, Charlie Y;Birdwell, L Dillon;Kohli, Rahul M
通讯作者：
Kohli, Rahul M

Nucleic acid determinants for selective deamination of DNA over RNA by activation-induced deaminase.

通过激活诱导的脱氨酶对 DNA 相对于 RNA 进行选择性脱氨的核酸决定簇。

DOI：
发表时间：
2013-08-27
期刊：
影响因子：
11.1
作者：
Nabel, Christopher S;Lee, Jae W;Wang, Laura C;Kohli, Rahul M
通讯作者：
Kohli, Rahul M

Introduction: Genome Modifying Mechanisms.

简介：基因组修饰机制。

DOI：
发表时间：
2016
期刊：
影响因子：
62.1
作者：
Kohli, Rahul M;Van Duyne, Gregory D
通讯作者：
Van Duyne, Gregory D

The expanding scope and impact of epigenetic cytosine modifications.

表观遗传胞嘧啶修饰的范围和影响不断扩大。

DOI：
10.1016/j.cbpa.2016.05.029
发表时间：
2016-08
期刊：
Current opinion in chemical biology
影响因子：
7.8
作者：
Liu MY;DeNizio JE;Schutsky EK;Kohli RM
通讯作者：
Kohli RM

Analysis of the machinery and intermediates of the 5 hmC ‐ mediated DNA demethylation pathway in aging on samples from the

老化样品中 5 hmC 介导的 DNA 去甲基化途径的机制和中间体分析

DOI：
10.1016/j.saa.2014.04.144
发表时间：
2024-09-13
期刊：
Spectrochimica acta. Part A, Molecular and biomolecular spectroscopy
影响因子：
0
作者：
Valentini;Zampieri;Malavolta;Bacalini;Roberta;V. Calabrese;Guastafierro;Reale;Franceschi;Hervonen;Koller;Bernhardt;Slagboom;Toussaint;K Sikora;Gonos;Breusing;Grune;Jansen;Dollé;Moreno;Villanueva;Sindlinger;Bürkle;Ciccarone;Paola;Caiafa
通讯作者：
Caiafa