Evolutionary Analysis and Comparative Genomics of Protein Superfamilies

蛋白质超家族的进化分析和比较基因组学

• 批准号: 8344972
• 负责人: Aravind Iyer
• 金额: $ 119.92万
• 依托单位: NATIONAL LIBRARY OF MEDICINE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8344972
• 关键词: Active Sites; Address; Adenylate Cyclase; Amino Acids; Amino Acyl-tRNA Synthetases; Anabolism; Architecture; Area; Bacteria; Berlin; Binding; Binding Sites; Biological; Biological Process; Biology; Catalysis; Catalytic Domain; Cations; Cells; Citric Acid Cycle; Classification; Collagen; Comparative Genomic Analysis; Computing Methodologies; Consult; Contractor; DNA Sequence Rearrangement; DNA Viruses; DNA-Directed DNA Polymerase; DNA-Directed RNA Polymerase; Dimerization; Dioxygenases; Disease; Dynamin; Enzymatic Biochemistry; Enzymes; Event; Evolution; Family; Ferredoxin; Genes; Genome; Glycine; Guanine Nucleotides; Guanosine Triphosphate; Guanosine Triphosphate Phosphohydrolases; Hydroxylation; Immunity; In Vitro; Intracellular Membranes; Iron; Isomerase; Journals; Life; Lipids; Lymphocyte; Manuscripts; Mediating; Membrane; Metabolism; Metals; Methods; Microscopic; Mixed Function Oxygenases; Modification; Molecular; Monograph; Mutagenesis; Nature; Neighborhoods; Nucleic Acids; Nucleotides; Pathway interactions; Pattern; Peer Review; Peptide Antibiotics; Peptides; Peroxidases; Phenylalanine-Specific tRNA; Play; Polymerase; Polynucleotide 5' -Hydroxyl-Kinase; Polynucleotides; Positioning Attribute; Postdoctoral Fellow; Process; Property; Proteins; Publications; Publishing; RNA; RNA-Directed DNA Polymerase; RNA-Directed RNA Polymerase; Radiation; Reaction; Recording of previous events; Research; Research Project Grants; Role; Science; Scientist; Siderophores; Signal Transduction; Specificity; Structure; Surface; System; Tertiary Protein Structure; Transcriptional Regulation; Transfer RNA; Work; alpha ketoglutarate; apoptosis in lymphocytes; base; comparative genomics; diguanylate cyclase; epimerase; genome sequencing; in vivo; innovation; inorganic phosphate; interest; member; novel; peroxidation; polymerization; programs; protein structure function; repaired; scaffold; sensor; sugar; translation factor; tripolyphosphate; viral RNA; wybutosine

Dr. Aravind has an ongoing interest in using computational methods to decipher various aspects of protein structure, function and evolution. During 2010, Dr. Aravind demonstrated exceptional progress and effective planning and execution of several major research projects along these lines. These research projects cover the areas of molecular enzymology, signal transduction and transcriptional regulation mechanisms using computational methods. His group comprising of 1 staff scientist, 3 post-doctoral fellows and one contractor has over 12 publications in peer-reviewed publications in top scientific journals. He also published a comprehensive monograph on signal sensor domains in bacteria, which is recognized as a major work in this field. In this period, Dr. Aravind was also consulted to serve as a referee for several manuscripts submitted to the journals Science, Cell, Genome Research, JMB and Nucleic Acids Research, Genome Biology. He was an invited to speaker at three venues in course of the year. Some highlights of Dr. Aravinds 2010 research program include the following: Dr. Aravinds group studied GTPases of immunity-associated proteins (GIMAPs), a distinctive family of GTPases, which control apoptosis in lymphocytes and play a central role in lymphocyte maturation and lymphocyte-associated diseases. To explore their function and mechanism, he collaborated with Olivier Daumkes group, Max-Delbrck-Centrum fr Molekulare Medizin, Berlin to determine crystal structures of a representative member, GIMAP2, in different nucleotide-loading and oligomerization states. Nucleotide-free and GDP-bound GIMAP2 were monomeric and revealed a guanine nucleotide-binding domain of the TRAFAC (translation factor associated) class with a unique amphipathic helix 7 packing against switch II. In the absence of 7 and the presence of GTP, GIMAP2 oligomerized via two distinct interfaces in the crystal. GTP-induced stabilization of switch I mediates dimerization across the nucleotide-binding site, which also involves the GIMAP specificity motif and the nucleotide base. Structural rearrangements in switch II appear to induce the release of 7 allowing oligomerization to proceed via a second interface. The unique architecture of the linear oligomer predicted by Dr. Aravinds analysis was confirmed by mutagenesis by Oliver Daumkes group. Furthermore, they showed a function for the GIMAP2 oligomer at the surface of lipid droplets. Although earlier studies indicated that GIMAPs are related to the septins, the current structure also revealed a strikingly similar nucleotide coordination and dimerization mode as in the dynamin GTPase. Based on this, Dr. Aravind reexamined the relationships of the septin- and dynamin-like GTPases and demonstrated that these are likely to have emerged from a common membrane-associated dimerizing ancestor. This ancestral property appears to be critical for the role of GIMAPs as nucleotide-regulated scaffolds on intracellular membranes. Dr. Aravind and his team performed from new analysis elucidating the structure and catalysis of a rather interesting novel class of nucleic acid polymerases. Almost all known nucleic acid polymerases catalyze 5'-3' polymerization by mediating the attack on an incoming nucleotide 5' triphosphate by the 3'OH from the growing polynucleotide chain in a template dependent or independent manner. The only known exception to this rule is the Thg1 RNA polymerase that catalyzes 3'-5' polymerization in vitro and also in vivo as a part of the maturation process of histidinyl tRNA. While the initial reaction catalyzed by Thg1 has been compared to adenylation catalyzed by the aminoacyl tRNA synthetases, the evolutionary relationships of Thg1 and the actual nature of the polymerase reaction catalyzed by it remain unclear. Using sensitive profile-profile comparison and structure prediction methods Dr. Aravind and his group showed that the catalytic domain Thg1 contains a RRM (ferredoxin) fold palm domain, just like the viral RNA-dependent RNA polymerases, reverse transcriptases, family A and B DNA polymerases, adenylyl cyclases, diguanylate cyclases (GGDEF domain) and the predicted polymerase of the CRISPR system. They showed just as in these polymerases, Thg1 possesses an active site with three acidic residues that chelate Mg++ cations. Based on this they predicted that Thg1 catalyzes polymerization similarly to the 5'-3' polymerases, but uses the incoming 3' OH to attack the 5' triphosphate generated at the end of the elongating polynucleotide. In addition they identified a distinct set of residues unique to Thg1 that they predicted as comprising a second active site, which catalyzes the initial adenylation reaction to prime 3'-5' polymerization. Based on contextual information from conserved gene neighborhoods they showed that Thg1 might function in conjunction with a polynucleotide kinase that generates an initial 5' phosphate substrate for it at the end of a RNA molecule. In addition to histidinyl tRNA maturation, Thg1 might have other RNA repair roles in representatives from all the three superkingdoms of life as well as certain large DNA viruses. They also present evidence that among the polymerase-like domains Thg1 is most closely related to the catalytic domains of the GGDEF and CRISPR polymerase proteins. Based on this relationship and the phyletic patterns of these enzymes they inferred that the Thg1 protein is likely to represent an archaeo-eukaryotic branch of the same clade of proteins that gave rise to the mobile CRISPR polymerases and in bacteria spawned the GGDEF domains. Thg1 is likely to be close to the ancestral version of this family of enzymes that might have played a role in RNA repair in the last universal common ancestor. A key work by Dr. Aravind and his group resulted in solving the evolutionary classification of the jumonji-like enzymes and predicting of the long elusive catalytic activity of the Wybutosine hydroxylase/peroxidase. Unlike classical 2-oxoglutarate and iron-dependent dioxygenases, which include several nucleic acid modifiers, the structurally similar jumonji-related dioxygenase superfamily was only known to catalyze peptide modifications. Using comparative genomics methods, Dr. Aravind and his team predicted that a family of jumonji-related enzymes catalyzes wybutosine hydroxylation/peroxidation at position 37 of eukaryotic tRNAPhe. Identification of this enzyme raised questions regarding the emergence of protein- and nucleic acid-modifying activities among jumonji-related domains. They addressed these with a natural classification of DSBH domains and reconstructed the precursor of the dioxygenases as a sugar-binding domain. This precursor gave rise to sugar epimerases and metal-binding sugar isomerases. The sugar isomerase active site was exapted for catalysis of oxygenation, with a radiation of these enzymes in bacteria, probably due to impetus from the primary oxygenation event in Earth's history. 2-Oxoglutarate-dependent versions appear to have further expanded with rise of the tricarboxylic acid cycle. They identified previously under-appreciated aspects of their active site and multiple independent innovations of 2-oxoacid-binding basic residues among these superfamilies. They showed that double-stranded -helix dioxygenases diversified extensively in biosynthesis and modification of halogenated siderophores, antibiotics, peptide secondary metabolites and glycine-rich collagen-like proteins in bacteria. Jumonji-related domains diversified into three distinct lineages in bacterial secondary metabolism systems and these were precursors of the three major clades of eukaryotic enzymes. The specificity of wybutosine hydroxylase/peroxidase probably relates to the structural similarity of the modified moiety to the ancestral amino acid substrate of this superfamily.

Aravind 博士一直对使用计算方法破译蛋白质结构、功能和进化的各个方面感兴趣。 2010 年，Aravind 博士在几个重大研究项目中取得了非凡的进展以及有效的规划和执行。这些研究项目涵盖使用计算方法的分子酶学、信号转导和转录调控机制领域。他的团队由 1 名科学家、3 名博士后研究员和 1 名承包商组成，在顶级科学期刊的同行评审出版物上发表了超过 12 篇论文。他还出版了一本关于细菌信号传感器领域的综合专着，被认为是该领域的重要著作。在此期间，Aravind博士还受邀担任投稿至《Science》、《Cell》、《Genome Research》、《JMB》和《Nucleic Acids Research》、《Genome Biology》等期刊的多篇稿件的审稿人。 年内，他受邀在三个场馆发表演讲。 Aravinds 博士 2010 年研究计划的一些亮点包括： Aravinds 博士小组研究了免疫相关蛋白 (GIMAP) 的 GTP 酶，这是一个独特的 GTP 酶家族，它控制淋巴细胞的凋亡，并在淋巴细胞成熟和淋巴细胞相关疾病中发挥核心作用。为了探索它们的功能和机制，他与柏林 Max-Delbrck-Centrum fr Molekulare Medizin 的 Olivier Daumkes 小组合作，确定了代表成员 GIMAP2 在不同核苷酸负载和寡聚状态下的晶体结构。无核苷酸且结合 GDP 的 GIMAP2 是单体，并揭示了 TRAFAC（翻译因子相关）类的鸟嘌呤核苷酸结合结构域，具有针对开关 II 的独特两亲性螺旋 7 包装。在 7 不存在且 GTP 存在的情况下，GIMAP2 通过晶体中两个不同的界面寡聚。 GTP 诱导的开关 I 稳定介导跨核苷酸结合位点的二聚化，这也涉及 GIMAP 特异性基序和核苷酸碱基。开关 II 中的结构重排似乎诱导了 7 的释放，从而允许寡聚化通过第二个界面进行。 Aravinds 博士分析预测的线性低聚物的独特结构通过 Oliver Daumkes 小组的诱变得到了证实。此外，他们还展示了 GIMAP2 寡聚物在脂滴表面的功能。尽管早期的研究表明 GIMAP 与脓蛋白有关，但目前的结构也揭示了与动力 GTP 酶中惊人相似的核苷酸配位和二聚化模式。在此基础上，Aravind 博士重新检查了类 Septin 和动力蛋白 GTP 酶的关系，并证明这些 GTP 酶很可能来自共同的膜相关二聚化祖先。这种祖先特性似乎对于 GIMAP 作为细胞内膜上核苷酸调节支架的作用至关重要。 Aravind 博士和他的团队进行了新的分析，阐明了一类相当有趣的新型核酸聚合酶的结构和催化作用。几乎所有已知的核酸聚合酶都通过介导来自生长的多核苷酸链的3'OH以模板依赖或独立的方式攻击进入的核苷酸5'三磷酸来催化5'-3'聚合。该规则唯一已知的例外是 Thg1 RNA 聚合酶，它在体外和体内催化 3'-5' 聚合，作为组氨酰 tRNA 成熟过程的一部分。虽然 Thg1 催化的初始反应与氨酰 tRNA 合成酶催化的腺苷酸化进行了比较，但 Thg1 的进化关系及其催化的聚合酶反应的实际性质仍不清楚。 Aravind 博士和他的团队使用灵敏的图谱比较和结构预测方法表明，催化结构域 Thg1 包含 RRM（铁氧化还原蛋白）折叠掌结构域，就像病毒 RNA 依赖性 RNA 聚合酶、逆转录酶、A 族和 B 族 DNA 一样聚合酶、腺苷酸环化酶、二鸟苷酸环化酶（GGDEF 结构域）和 CRISPR 系统的预测聚合酶。他们表明，就像在这些聚合酶中一样，Thg1 拥有一个带有三个酸性残基的活性位点，可以螯合 Mg++ 阳离子。基于此，他们预测Thg1与5'-3'聚合酶类似地催化聚合，但使用引入的3'OH来攻击在延伸多核苷酸末端产生的5'三磷酸。此外，他们还确定了 Thg1 特有的一组独特残基，他们预测这些残基包含第二个活性位点，可催化最初的腺苷酸化反应引发 3'-5' 聚合。根据保守基因邻域的背景信息，他们表明 Thg1 可能与多核苷酸激酶结合发挥作用，该激酶在 RNA 分子末端为其生成初始 5' 磷酸底物。除了组氨酰 tRNA 成熟之外，Thg1 可能在所有三个生命超级王国以及某些大型 DNA 病毒的代表中具有其他 RNA 修复作用。他们还提供了证据，表明在聚合酶样结构域中，Thg1 与 GGDEF 和 CRISPR 聚合酶蛋白的催化结构域关系最为密切。基于这种关系和这些酶的系统发育模式，他们推断 Thg1 蛋白很可能代表同一蛋白质进化枝的古真核分支，该蛋白质进化枝产生了移动 CRISPR 聚合酶，并在细菌中产生了 GGDEF 结构域。 Thg1 很可能接近该酶家族的祖先版本，该酶家族可能在最后一个普遍共同祖先的 RNA 修复中发挥了作用。 Aravind 博士和他的团队的一项关键工作解决了十文字类酶的进化分类问题，并预测了威布托辛羟化酶/过氧化物酶长期难以捉摸的催化活性。与包含多种核酸修饰剂的经典 2-酮戊二酸和铁依赖性双加氧酶不同，结构相似的 jumonji 相关双加氧酶超家族仅已知能催化肽修饰。 Aravind 博士和他的团队利用比较基因组学方法预测，Jumonji 相关酶家族会催化真核生物 tRNAPhe 37 位的威布托辛羟基化/过氧化反应。这种酶的鉴定引发了关于十文字相关域中蛋白质和核酸修饰活性的出现的问题。他们通过 DSBH 结构域的自然分类来解决这些问题，并将双加氧酶的前体重建为糖结合结构域。该前体产生糖差向异构酶和金属结合糖异构酶。糖异构酶活性位点适合催化氧化，这些酶在细菌中辐射，可能是由于地球历史上初次氧化事件的推动。随着三羧酸循环的兴起，2-氧戊二酸依赖性形式似乎进一步扩展。他们确定了其活性位点的先前未被充分认识的方面以及这些超家族中 2-含氧酸结合碱性残基的多项独立创新。他们表明，双链β-螺旋双加氧酶在细菌中卤化铁载体、抗生素、肽次级代谢产物和富含甘氨酸的胶原蛋白样蛋白的生物合成和修饰中广泛多样化。十文字相关结构域在细菌次生代谢系统中分化成三个不同的谱系，它们是真核酶三个主要分支的前体。威布托辛羟化酶/过氧化物酶的特异性可能与修饰部分与该超家族的祖先氨基酸底物的结构相似性有关。