Stoichiometry of ligand binding and role of C-terminal lysines in Mycobacterium tuberculosis and human GAPDH multifunctionality.

Ajay Kumar, Rajender Kumar, Vishant Mahendra Boradia, Himanshu Malhotra, Adarsh Kumar, Sriraj Seth, Prabha Garg, Subramanian Karthikeyan, Manoj Raje, Chaaya Iyengar Raje
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Abstract

Glyceraldehyde-3-phosphate-dehydrogenase (GAPDH; EC1.2.1.12) has several functions in Mycobacterium tuberculosis (Mtb) and the human host. Apart from its role in glycolysis, it serves both as a cell surface and a secreted receptor for plasmin(ogen) (Plg/Plm), transferrin (Tf), and lactoferrin (Lf). Plg sequestration by Mtb GAPDH facilitates bacterial adhesion and tissue invasion, while an equivalent interaction with host GAPDH regulates immune cell migration. In both, host and microbe, internalization of Tf/Lf-GAPDH complexes serves as a route for iron acquisition. To date, the structure of Mtb GAPDH or the residues involved in these moonlighting interactions have not been identified. This study provides the first known X-ray crystal structure of Mtb GAPDH. Through further mutagenesis and functional assays, we found that the C-terminal lysines of Mtb and human GAPDH affect enzyme activity and ligand binding. We also establish the stoichiometry of Plg, Tf and Lf interactions with the GAPDH tetramer. Lastly, molecular simulation studies reveal the interactions of the C-terminal lysine residues.

配体结合的化学计量以及 C 端赖氨酸在结核分枝杆菌和人类 GAPDH 多功能性中的作用。
甘油醛-3-磷酸脱氢酶(GAPDH;EC1.2.1.12)在结核分枝杆菌(Mtb)和人类宿主体内具有多种功能。除了在糖酵解中的作用外,它还是plasmin(ogen)(Plg/Plm)、转铁蛋白(Tf)和乳铁蛋白(Lf)的细胞表面和分泌受体。Plg 被 Mtb GAPDH 封闭可促进细菌粘附和组织侵袭,而与宿主 GAPDH 的等效相互作用可调节免疫细胞的迁移。在宿主和微生物中,Tf/Lf-GAPDH 复合物的内化都是获取铁的途径。迄今为止,Mtb GAPDH 的结构或参与这些月光相互作用的残基尚未确定。本研究首次提供了已知的 Mtb GAPDH 的 X 射线晶体结构。通过进一步的诱变和功能测试,我们发现 Mtb 和人类 GAPDH 的 C 端赖氨酸会影响酶活性和配体结合。我们还确定了 Plg、Tf 和 Lf 与 GAPDH 四聚体相互作用的比例。最后,分子模拟研究揭示了 C 端赖氨酸残基的相互作用。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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