The Crystal Structure of Nα-p-tosyl-lysyl Chloromethylketone-Bound Oligopeptidase B from Serratia Proteamaculans Revealed a New Type of Inhibitor Binding

A covalent serine protease inhibitor—Na-p-Tosyl-Lysyl Chloromethylketone (TCK) is a modified lysine residue tosylated at the N-terminus and chloromethylated at the C-terminus, one molecule of which is capable of forming two covalent bonds with both Ser and His catalytic residues, was co-crystallized...

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Autores principales:	Vladimir I. Timofeev, Dmitry E. Petrenko, Yulia K. Agapova, Anna V. Vlaskina, David M. Karlinsky, Anna G. Mikhailova, Inna P. Kuranova, Tatiana V. Rakitina
Formato:	article
Lenguaje:	EN
Publicado:	MDPI AG 2021
Materias:	prolyloligopeptidase oligopeptidase B crystal structure intermediate state serine protease chloromethylketone inhibitors Crystallography QD901-999
Acceso en línea:	https://doaj.org/article/7504dffdaa2e4f73933f2fbd6cbe21c1
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spelling	oai:doaj.org-article:7504dffdaa2e4f73933f2fbd6cbe21c12021-11-25T17:19:47ZThe Crystal Structure of Nα-p-tosyl-lysyl Chloromethylketone-Bound Oligopeptidase B from Serratia Proteamaculans Revealed a New Type of Inhibitor Binding10.3390/cryst111114382073-4352https://doaj.org/article/7504dffdaa2e4f73933f2fbd6cbe21c12021-11-01T00:00:00Zhttps://www.mdpi.com/2073-4352/11/11/1438https://doaj.org/toc/2073-4352A covalent serine protease inhibitor—Na-p-Tosyl-Lysyl Chloromethylketone (TCK) is a modified lysine residue tosylated at the N-terminus and chloromethylated at the C-terminus, one molecule of which is capable of forming two covalent bonds with both Ser and His catalytic residues, was co-crystallized with modified oligopeptidase B (OpB) from <i>Serratia proteomaculans</i> (PSPmod). The kinetics study, which preceded crystallization, shows that the stoichiometry of TCK-dependent inhibition of PSPmod was 1:2 (protein:inhibitor). The crystal structure of the PSPmod-TCK complex, solved at a resolution of 2.3 Å, confirmed a new type of inhibitor binding. Two TCK molecules were bound to one enzyme molecule: one with the catalytic Ser, the other with the catalytic His. Due to this mode of binding, the intermediate state of PSPmod and the disturbed conformation of the catalytic triad were preserved in the PSPmod-TCK complex. Nevertheless, the analysis of the amino acid surroundings of the inhibitor molecule bound to the catalytic Ser and its comparison with that of antipain-bound OpB from <i>Trypanosoma brucei</i> provided an insight in the structure of the PSPmod substrate-binding pocket. Supposedly, the new type of binding is typical for the interaction of chloromethylketone derivatives with two-domain OpBs. In the open conformational state that these enzymes are assumed in solution, the disordered configuration of the catalytic triad prevents simultaneous interaction of one inhibitor molecule with two catalytic residues.Vladimir I. TimofeevDmitry E. PetrenkoYulia K. AgapovaAnna V. VlaskinaDavid M. KarlinskyAnna G. MikhailovaInna P. KuranovaTatiana V. RakitinaMDPI AGarticleprolyloligopeptidaseoligopeptidase Bcrystal structureintermediate stateserine proteasechloromethylketone inhibitorsCrystallographyQD901-999ENCrystals, Vol 11, Iss 1438, p 1438 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	prolyloligopeptidase oligopeptidase B crystal structure intermediate state serine protease chloromethylketone inhibitors Crystallography QD901-999
spellingShingle	prolyloligopeptidase oligopeptidase B crystal structure intermediate state serine protease chloromethylketone inhibitors Crystallography QD901-999 Vladimir I. Timofeev Dmitry E. Petrenko Yulia K. Agapova Anna V. Vlaskina David M. Karlinsky Anna G. Mikhailova Inna P. Kuranova Tatiana V. Rakitina The Crystal Structure of Nα-p-tosyl-lysyl Chloromethylketone-Bound Oligopeptidase B from Serratia Proteamaculans Revealed a New Type of Inhibitor Binding
description	A covalent serine protease inhibitor—Na-p-Tosyl-Lysyl Chloromethylketone (TCK) is a modified lysine residue tosylated at the N-terminus and chloromethylated at the C-terminus, one molecule of which is capable of forming two covalent bonds with both Ser and His catalytic residues, was co-crystallized with modified oligopeptidase B (OpB) from <i>Serratia proteomaculans</i> (PSPmod). The kinetics study, which preceded crystallization, shows that the stoichiometry of TCK-dependent inhibition of PSPmod was 1:2 (protein:inhibitor). The crystal structure of the PSPmod-TCK complex, solved at a resolution of 2.3 Å, confirmed a new type of inhibitor binding. Two TCK molecules were bound to one enzyme molecule: one with the catalytic Ser, the other with the catalytic His. Due to this mode of binding, the intermediate state of PSPmod and the disturbed conformation of the catalytic triad were preserved in the PSPmod-TCK complex. Nevertheless, the analysis of the amino acid surroundings of the inhibitor molecule bound to the catalytic Ser and its comparison with that of antipain-bound OpB from <i>Trypanosoma brucei</i> provided an insight in the structure of the PSPmod substrate-binding pocket. Supposedly, the new type of binding is typical for the interaction of chloromethylketone derivatives with two-domain OpBs. In the open conformational state that these enzymes are assumed in solution, the disordered configuration of the catalytic triad prevents simultaneous interaction of one inhibitor molecule with two catalytic residues.
format	article
author	Vladimir I. Timofeev Dmitry E. Petrenko Yulia K. Agapova Anna V. Vlaskina David M. Karlinsky Anna G. Mikhailova Inna P. Kuranova Tatiana V. Rakitina
author_facet	Vladimir I. Timofeev Dmitry E. Petrenko Yulia K. Agapova Anna V. Vlaskina David M. Karlinsky Anna G. Mikhailova Inna P. Kuranova Tatiana V. Rakitina
author_sort	Vladimir I. Timofeev
title	The Crystal Structure of Nα-p-tosyl-lysyl Chloromethylketone-Bound Oligopeptidase B from Serratia Proteamaculans Revealed a New Type of Inhibitor Binding
title_short	The Crystal Structure of Nα-p-tosyl-lysyl Chloromethylketone-Bound Oligopeptidase B from Serratia Proteamaculans Revealed a New Type of Inhibitor Binding
title_full	The Crystal Structure of Nα-p-tosyl-lysyl Chloromethylketone-Bound Oligopeptidase B from Serratia Proteamaculans Revealed a New Type of Inhibitor Binding
title_fullStr	The Crystal Structure of Nα-p-tosyl-lysyl Chloromethylketone-Bound Oligopeptidase B from Serratia Proteamaculans Revealed a New Type of Inhibitor Binding
title_full_unstemmed	The Crystal Structure of Nα-p-tosyl-lysyl Chloromethylketone-Bound Oligopeptidase B from Serratia Proteamaculans Revealed a New Type of Inhibitor Binding
title_sort	crystal structure of nα-p-tosyl-lysyl chloromethylketone-bound oligopeptidase b from serratia proteamaculans revealed a new type of inhibitor binding
publisher	MDPI AG
publishDate	2021
url	https://doaj.org/article/7504dffdaa2e4f73933f2fbd6cbe21c1
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The Crystal Structure of Nα-p-tosyl-lysyl Chloromethylketone-Bound Oligopeptidase B from Serratia Proteamaculans Revealed a New Type of Inhibitor Binding

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