Structure of human factor VIIa–soluble tissue factor with calcium, magnesium and rubidium

Autor:	Kaillathe Padmanabhan, Sriram Krishnaswamy, Hans Brandstetter, Amy E. Schmidt, Duilio Cascio, S. Paul Bajaj, Kanagasabai Vadivel
Rok vydání:	2021
Předmět:	medicine.medical_treatment Sodium chemistry.chemical_element Factor VIIa Calcium 01 natural sciences Rubidium Structure-Activity Relationship 03 medical and health sciences Tissue factor Thrombin Protein Domains Structural Biology TheoryofComputation_ANALYSISOFALGORITHMSANDPROBLEMCOMPLEXITY 0103 physical sciences medicine Humans Magnesium 030304 developmental biology Gla domain 0303 health sciences Binding Sites Protease 010304 chemical physics Chemistry Substrate (chemistry) Research Papers Recombinant Proteins Crystallography Protein Binding medicine.drug
Zdroj:	Acta Crystallogr D Struct Biol
ISSN:	2059-7983
Popis:	Coagulation factor VIIa (FVIIa) consists of a γ-carboxyglutamic acid (GLA) domain, two epidermal growth factor-like (EGF) domains and a protease domain. FVIIa binds three Mg2+ ions and four Ca2+ ions in the GLA domain, one Ca2+ ion in the EGF1 domain and one Ca2+ ion in the protease domain. Further, FVIIa contains an Na+ site in the protease domain. Since Na+ and water share the same number of electrons, Na+ sites in proteins are difficult to distinguish from waters in X-ray structures. Here, to verify the Na+ site in FVIIa, the structure of the FVIIa–soluble tissue factor (TF) complex was solved at 1.8 Å resolution containing Mg2+, Ca2+ and Rb+ ions. In this structure, Rb+ replaced two Ca2+ sites in the GLA domain and occupied three non-metal sites in the protease domain. However, Rb+ was not detected at the expected Na+ site. In kinetic experiments, Na+ increased the amidolytic activity of FVIIa towards the synthetic substrate S-2288 (H-D-Ile-Pro-Arg-p-nitroanilide) by ∼20-fold; however, in the presence of Ca2+, Na+ had a negligible effect. Ca2+ increased the hydrolytic activity of FVIIa towards S-2288 by ∼60-fold in the absence of Na+ and by ∼82-fold in the presence of Na+. In molecular-dynamics simulations, Na+ stabilized the two Na+-binding loops (the 184-loop and 220-loop) and the TF-binding region spanning residues 163–180. Ca2+ stabilized the Ca2+-binding loop (the 70-loop) and Na+-binding loops but not the TF-binding region. Na+ and Ca2+ together stabilized both the Na+-binding and Ca2+-binding loops and the TF-binding region. Previously, Rb+ has been used to define the Na+ site in thrombin; however, it was unsuccessful in detecting the Na+ site in FVIIa. A conceivable explanation for this observation is provided.
Databáze:	OpenAIRE
Externí odkaz:	https://explore.openaire.eu/search/publication?articleId=doi_dedup___::768553e62fbb6b3b4a9e8f6a22f86808 https://doi.org/10.1107/s2059798321003922 Zobrazit plný text záznamu Plný text ve formátu PDF Plný text ve formátu HTML
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