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[摘要]：Factor XIII catalyzes formation of gamma-glutamyl-epsilon-lysyl crosslinks within fibrin clots. FXIII A(2) can be activated proteolytically with thrombin and low mM Ca2+ or nonproteolytically with high monovalent/divalent cations along with low mM Ca2+. Physiologically, FXIII A(2) is poised to respond to transient influxes of Ca2+ in a Na+ containing environment. A successful strategy to monitor FXIII conformational events is hydrogen-deuterium exchange (HDX) coupled with mass spectrometry. FXIII A(2) was examined in the presence of different cations (Ca2+, Mg2+, Ba2+, Cu2+, Na+, TMAC(+), and EDA(2+)) ranging from 1 to 2 mM, physiological Ca2+ concentration, to 50-500 mM for nonproteolytic activation. Increases in FXIII solvent exposure could already be observed at 1 mM Ca2+ for the dimer interface, the catalytic site, and glutamine substrate regions. By contrast, solvent protection was observed at the secondary cleavage site. These events occurred even though 1 mM Ca2+ is insufficient for FXIII activation. The metals 1 mM Mg2+, 1 mM Ba2+, and 1 mM Cu2+ each led to conformational changes, many in the same FXIII regions as Ca2+. FXIII could also be activated nonproteolytically with 500 mM tetramethylammonium chloride (TMAC(+)) and 500 mM ethylenediamine (EDA(2+)), both with 2 mM Ca2+. These different HDX studies help reveal the first FXIII segments that respond to physiological Ca2+ levels. (c) 2011 Elsevier Inc. All rights reserved. |
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