Serine protease dynamics revealed by NMR analysis of the thrombin-thrombomodulin complex

Abstract Serine proteases catalyze a multi-step covalent catalytic mechanism of peptide bond cleavage. It has long been assumed that serine proteases including thrombin carry-out catalysis without significant conformational rearrangement of their stable two-β-barrel structure. We present nuclear mag...

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Autores principales:	Riley B. Peacock, Taylor McGrann, Marco Tonelli, Elizabeth A. Komives
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/88183b8ffe2e4f00a14e25f6f59fd4a8
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spelling	oai:doaj.org-article:88183b8ffe2e4f00a14e25f6f59fd4a82021-12-02T17:39:20ZSerine protease dynamics revealed by NMR analysis of the thrombin-thrombomodulin complex10.1038/s41598-021-88432-z2045-2322https://doaj.org/article/88183b8ffe2e4f00a14e25f6f59fd4a82021-04-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-88432-zhttps://doaj.org/toc/2045-2322Abstract Serine proteases catalyze a multi-step covalent catalytic mechanism of peptide bond cleavage. It has long been assumed that serine proteases including thrombin carry-out catalysis without significant conformational rearrangement of their stable two-β-barrel structure. We present nuclear magnetic resonance (NMR) and hydrogen deuterium exchange mass spectrometry (HDX-MS) experiments on the thrombin-thrombomodulin (TM) complex. Thrombin promotes procoagulative fibrinogen cleavage when fibrinogen engages both the anion binding exosite 1 (ABE1) and the active site. It is thought that TM promotes cleavage of protein C by engaging ABE1 in a similar manner as fibrinogen. Thus, the thrombin-TM complex may represent the catalytically active, ABE1-engaged thrombin. Compared to apo- and active site inhibited-thrombin, we show that thrombin-TM has reduced μs-ms dynamics in the substrate binding (S1) pocket consistent with its known acceleration of protein C binding. Thrombin-TM has increased μs-ms dynamics in a β-strand connecting the TM binding site to the catalytic aspartate. Finally, thrombin-TM had doublet peaks indicative of dynamics that are slow on the NMR timescale in residues along the interface between the two β-barrels. Such dynamics may be responsible for facilitating the N-terminal product release and water molecule entry that are required for hydrolysis of the acyl-enzyme intermediate.Riley B. PeacockTaylor McGrannMarco TonelliElizabeth A. KomivesNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-13 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Riley B. Peacock Taylor McGrann Marco Tonelli Elizabeth A. Komives Serine protease dynamics revealed by NMR analysis of the thrombin-thrombomodulin complex
description	Abstract Serine proteases catalyze a multi-step covalent catalytic mechanism of peptide bond cleavage. It has long been assumed that serine proteases including thrombin carry-out catalysis without significant conformational rearrangement of their stable two-β-barrel structure. We present nuclear magnetic resonance (NMR) and hydrogen deuterium exchange mass spectrometry (HDX-MS) experiments on the thrombin-thrombomodulin (TM) complex. Thrombin promotes procoagulative fibrinogen cleavage when fibrinogen engages both the anion binding exosite 1 (ABE1) and the active site. It is thought that TM promotes cleavage of protein C by engaging ABE1 in a similar manner as fibrinogen. Thus, the thrombin-TM complex may represent the catalytically active, ABE1-engaged thrombin. Compared to apo- and active site inhibited-thrombin, we show that thrombin-TM has reduced μs-ms dynamics in the substrate binding (S1) pocket consistent with its known acceleration of protein C binding. Thrombin-TM has increased μs-ms dynamics in a β-strand connecting the TM binding site to the catalytic aspartate. Finally, thrombin-TM had doublet peaks indicative of dynamics that are slow on the NMR timescale in residues along the interface between the two β-barrels. Such dynamics may be responsible for facilitating the N-terminal product release and water molecule entry that are required for hydrolysis of the acyl-enzyme intermediate.
format	article
author	Riley B. Peacock Taylor McGrann Marco Tonelli Elizabeth A. Komives
author_facet	Riley B. Peacock Taylor McGrann Marco Tonelli Elizabeth A. Komives
author_sort	Riley B. Peacock
title	Serine protease dynamics revealed by NMR analysis of the thrombin-thrombomodulin complex
title_short	Serine protease dynamics revealed by NMR analysis of the thrombin-thrombomodulin complex
title_full	Serine protease dynamics revealed by NMR analysis of the thrombin-thrombomodulin complex
title_fullStr	Serine protease dynamics revealed by NMR analysis of the thrombin-thrombomodulin complex
title_full_unstemmed	Serine protease dynamics revealed by NMR analysis of the thrombin-thrombomodulin complex
title_sort	serine protease dynamics revealed by nmr analysis of the thrombin-thrombomodulin complex
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/88183b8ffe2e4f00a14e25f6f59fd4a8
work_keys_str_mv	AT rileybpeacock serineproteasedynamicsrevealedbynmranalysisofthethrombinthrombomodulincomplex AT taylormcgrann serineproteasedynamicsrevealedbynmranalysisofthethrombinthrombomodulincomplex AT marcotonelli serineproteasedynamicsrevealedbynmranalysisofthethrombinthrombomodulincomplex AT elizabethakomives serineproteasedynamicsrevealedbynmranalysisofthethrombinthrombomodulincomplex
_version_	1718379833817300992

Serine protease dynamics revealed by NMR analysis of the thrombin-thrombomodulin complex

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