| Autor: |
Svirid AV; Institute of Bioorganic Chemistry, National Academy of Sciences of Belarus, Akad. Kuprevicha Str. 5 /2, Minsk, 220141, Belarus., Ershov PV; Institute of Biomedical Chemistry, Pogodinskaya Street 10, bldg. 8, Moscow, 119121, Russia., Yablokov EO; Institute of Biomedical Chemistry, Pogodinskaya Street 10, bldg. 8, Moscow, 119121, Russia., Kaluzhskiy LA; Institute of Biomedical Chemistry, Pogodinskaya Street 10, bldg. 8, Moscow, 119121, Russia., Mezentsev YV; Institute of Biomedical Chemistry, Pogodinskaya Street 10, bldg. 8, Moscow, 119121, Russia., Florinskaya AV; Institute of Biomedical Chemistry, Pogodinskaya Street 10, bldg. 8, Moscow, 119121, Russia., Sushko TA; Institute of Bioorganic Chemistry, National Academy of Sciences of Belarus, Akad. Kuprevicha Str. 5 /2, Minsk, 220141, Belarus.; Department of Bioengineering, School of Engineering, The University of Tokyo, Tokyo, 108-8639, Japan., Strushkevich NV; Institute of Bioorganic Chemistry, National Academy of Sciences of Belarus, Akad. Kuprevicha Str. 5 /2, Minsk, 220141, Belarus., Gilep AA; Institute of Bioorganic Chemistry, National Academy of Sciences of Belarus, Akad. Kuprevicha Str. 5 /2, Minsk, 220141, Belarus., Usanov SA; Institute of Bioorganic Chemistry, National Academy of Sciences of Belarus, Akad. Kuprevicha Str. 5 /2, Minsk, 220141, Belarus., Medvedev AE; Institute of Biomedical Chemistry, Pogodinskaya Street 10, bldg. 8, Moscow, 119121, Russia., Ivanov AS; Institute of Biomedical Chemistry, Pogodinskaya Street 10, bldg. 8, Moscow, 119121, Russia. |
| Abstrakt: |
Thromboxane synthase (TBXAS1) catalyzes the isomerization reaction of prostaglandin H2 producing thromboxane A2, the autocrine and paracrine factor in many cell types. A high activity and metastability by these arachidonic acid derivatives suggests the existence of supramolecular structures that are involved in the regulation of the biosynthesis and directed translocation of thromboxane to the receptor. The objective of this study was to identify TBXAS1 protein partners from human liver tissue lysate using a complex approach based on the direct molecular fishing technique, LC-MS/MS protein identification, and protein-protein interaction validation by surface plasmon resonance (SPR). As a result, 12 potential TBXAS1 protein partners were identified, including the components regulating cytoskeleton organization (BBIP1 and ANKMY1), components of the coagulation cascade of human blood (SERPINA1, SERPINA3, APOH, FGA, and FN1), and the enzyme involved in the metabolism of xenobiotics and endogenous bioregulators (CYP2E1). SPR validation on the Biacore 3000 biosensor confirmed the effectiveness of the interaction between CYP2E1 (the enzyme that converts prostaglandin H2 to 12-HHT/thromboxane A2 proantagonist) and TBXAS1 ( K d = (4.3 ± 0.4) × 10 -7 M). Importantly, the TBXAS1•CYP2E1 complex formation increases fivefold in the presence of isatin (indole-2,3-dione, a low-molecular nonpeptide endogenous bioregulator, a product of CYP2E1). These results suggest that the interaction between these hemoproteins is important in the regulation of the biosynthesis of eicosanoids. |
