Pasquali Camila Cristina, Islam Zeyaul, Adamoski Douglas, Ferreira Igor Monteze, Righeto Ricardo Diogo, Bettini Jefferson, Portugal Rodrigo Villares, Yue Wyatt Wai-Yin, Gonzalez Ana, Dias Sandra Martha Gomes, Ambrosio Andre Luis Berteli
From the Laboratório Nacional de Biociências, Centro Nacional de Pesquisa em Energia e Materiais, Campinas, São Paulo 13083-970, Brazil.
the Structural Genomics Consortium, University of Oxford, Oxford OX3 7DQ, United Kingdom.
J Biol Chem. 2017 Jul 7;292(27):11572-11585. doi: 10.1074/jbc.M117.787291. Epub 2017 May 19.
On the basis of tissue-specific enzyme activity and inhibition by catalytic products, Hans Krebs first demonstrated the existence of multiple glutaminases in mammals. Currently, two human genes are known to encode at least four glutaminase isoforms. However, the phylogeny of these medically relevant enzymes remains unclear, prompting us to investigate their origin and evolution. Using prokaryotic and eukaryotic glutaminase sequences, we built a phylogenetic tree whose topology suggested that the multidomain architecture was inherited from bacterial ancestors, probably simultaneously with the hosting of the proto-mitochondrion endosymbiont. We propose an evolutionary model wherein the appearance of the most active enzyme isoform, glutaminase C (GAC), which is expressed in many cancers, was a late retrotransposition event that occurred in fishes from the Chondrichthyes class. The ankyrin (ANK) repeats in the glutaminases were acquired early in their evolution. To obtain information on ANK folding, we solved two high-resolution structures of the ANK repeat-containing C termini of both kidney-type glutaminase (KGA) and GLS2 isoforms (glutaminase B and liver-type glutaminase). We found that the glutaminase ANK repeats form unique intramolecular contacts through two highly conserved motifs; curiously, this arrangement occludes a region usually involved in ANK-mediated protein-protein interactions. We also solved the crystal structure of full-length KGA and present a small-angle X-ray scattering model for full-length GLS2. These structures explain these proteins' compromised ability to assemble into catalytically active supra-tetrameric filaments, as previously shown for GAC. Collectively, these results provide information about glutaminases that may aid in the design of isoform-specific glutaminase inhibitors.
基于组织特异性酶活性以及催化产物的抑制作用，汉斯·克雷布斯首次证明了哺乳动物中存在多种谷氨酰胺酶。目前，已知两个人类基因可编码至少四种谷氨酰胺酶同工型。然而，这些与医学相关的酶的系统发育仍不清楚，这促使我们对它们的起源和进化进行研究。利用原核生物和真核生物的谷氨酰胺酶序列，我们构建了一个系统发育树，其拓扑结构表明多结构域架构是从细菌祖先遗传而来的，可能与原线粒体共生体的宿主化同时发生。我们提出了一种进化模型，其中在许多癌症中表达的最活跃的酶同工型谷氨酰胺酶C（GAC）的出现是软骨鱼类鱼类中发生的晚期逆转座事件。谷氨酰胺酶中的锚蛋白（ANK）重复序列在其进化早期就已获得。为了获得有关ANK折叠的信息，我们解析了肾型谷氨酰胺酶（KGA）和GLS2同工型（谷氨酰胺酶B和肝型谷氨酰胺酶）含ANK重复序列的C末端的两个高分辨率结构。我们发现谷氨酰胺酶ANK重复序列通过两个高度保守的基序形成独特的分子内接触；奇怪的是，这种排列方式封闭了通常参与ANK介导的蛋白质-蛋白质相互作用的区域。我们还解析了全长KGA的晶体结构，并提出了全长GLS2的小角X射线散射模型。这些结构解释了这些蛋白质组装成具有催化活性的超四聚体细丝的能力受损，如先前对GAC的研究所示。总的来说，这些结果提供了有关谷氨酰胺酶的信息，可能有助于设计同工型特异性谷氨酰胺酶抑制剂。
