Molecular recognition in gas phase: theoretical and experimental study of non-covalent protein-ligand complexes by mass-spectrometry

In the present thesis we have explored different factors that impede accurate quantitative description of non-covalent protein-protein and protein-ligand interactions and design of new potent and specific binders from the scratch. Firstly, we addressed the role of solvent in the mechanism of non-cov...

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Detalhes bibliográficos
Autor: Dyachenko, Andrey
Tipo de documento: tese
Estado:Versão publicada
Data de publicação:2013
País:España
Recursos:CBUC, CESCA
Repositório:TDR. Tesis Doctorales en Red
OAI Identifier:oai:www.tdx.cat:10803/113301
Acesso em linha:http://hdl.handle.net/10803/113301
Access Level:Acceso aberto
Palavra-chave:Espectrometria de masses
Espectrometría de masas
Mass spectrometry
Interaccions no covalents
Interacciones no covalentes
Non-covalent interactions
Interaccions proteïna-proteïna
Interacciones proteína-proteína
Protein-protein interactions
Interaccions proteïna-lligand
Interacciones proteína-ligando
Ligand-protein interactions
Ciències Experimentals i Matemàtiques
547
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oai_identifier_str oai:www.tdx.cat:10803/113301
network_acronym_str ES
network_name_str España
repository_id_str
dc.title.none.fl_str_mv Molecular recognition in gas phase: theoretical and experimental study of non-covalent protein-ligand complexes by mass-spectrometry
title Molecular recognition in gas phase: theoretical and experimental study of non-covalent protein-ligand complexes by mass-spectrometry
spellingShingle Molecular recognition in gas phase: theoretical and experimental study of non-covalent protein-ligand complexes by mass-spectrometry
Dyachenko, Andrey
Espectrometria de masses
Espectrometría de masas
Mass spectrometry
Interaccions no covalents
Interacciones no covalentes
Non-covalent interactions
Interaccions proteïna-proteïna
Interacciones proteína-proteína
Protein-protein interactions
Interaccions proteïna-lligand
Interacciones proteína-ligando
Ligand-protein interactions
Ciències Experimentals i Matemàtiques
547
title_short Molecular recognition in gas phase: theoretical and experimental study of non-covalent protein-ligand complexes by mass-spectrometry
title_full Molecular recognition in gas phase: theoretical and experimental study of non-covalent protein-ligand complexes by mass-spectrometry
title_fullStr Molecular recognition in gas phase: theoretical and experimental study of non-covalent protein-ligand complexes by mass-spectrometry
title_full_unstemmed Molecular recognition in gas phase: theoretical and experimental study of non-covalent protein-ligand complexes by mass-spectrometry
title_sort Molecular recognition in gas phase: theoretical and experimental study of non-covalent protein-ligand complexes by mass-spectrometry
dc.creator.none.fl_str_mv Dyachenko, Andrey
author Dyachenko, Andrey
author_facet Dyachenko, Andrey
author_role author
dc.contributor.none.fl_str_mv Giralt Lledó, Ernest
Universitat de Barcelona. Departament de Química Orgànica
dc.subject.none.fl_str_mv Espectrometria de masses
Espectrometría de masas
Mass spectrometry
Interaccions no covalents
Interacciones no covalentes
Non-covalent interactions
Interaccions proteïna-proteïna
Interacciones proteína-proteína
Protein-protein interactions
Interaccions proteïna-lligand
Interacciones proteína-ligando
Ligand-protein interactions
Ciències Experimentals i Matemàtiques
547
topic Espectrometria de masses
Espectrometría de masas
Mass spectrometry
Interaccions no covalents
Interacciones no covalentes
Non-covalent interactions
Interaccions proteïna-proteïna
Interacciones proteína-proteína
Protein-protein interactions
Interaccions proteïna-lligand
Interacciones proteína-ligando
Ligand-protein interactions
Ciències Experimentals i Matemàtiques
547
description In the present thesis we have explored different factors that impede accurate quantitative description of non-covalent protein-protein and protein-ligand interactions and design of new potent and specific binders from the scratch. Firstly, we addressed the role of solvent in the mechanism of non-covalent interactions. Secondly, we tackled the question about the intrinsic conformational flexibility of the protein molecules and the part it plays in weak interactions between proteins. In the first part of the thesis we studied the interactions of vascular endothelial growth factor (VEGF) protein with five cyclic peptides in solution and gas phase. The results showed that affinities of five ligands to VEGF in solution and gas phase are ranked in inversed order. That is, the that has the highest affinity in solution (as shown by chemical shift perturbation NMR and isothermal titration calorimetry) forms the weakest complex with VEGF in gas phase, and vice versa. We compared gas-phase and solution binding affinities of of five peptides and made qualitative conclusions about the role of the solvent in protein-ligand interactions. In order to obtain more quantitative information about the gas-phase behavior of non-covalent complexes we have developed a combined experimental/theoretical approach to study the energetics of collisional activation of the ion prior to dissociation. We applied developed strategy to model CID in traveling wave ion guide (TWIG) collision cell. We validated the model on the CID of leu-enkephalin peptide and then applied developed strategy to five non-covalent protein-peptide complexes and found activation energies of their dissociation reactions. Next we applied ESI native MS to study the allosteric interactions between the molecular chaperonin GroEL and ATP. The obtained data allowed to construct a scheme of conformational transition of GroEL upon binding of ATP and distinguish between two different cooperativity models, providing strong arguments in favor of Monod-Wyman-Changeux (MWC) model. Finally, be studied the backbone dynamics of VEGF with a combination of NMR relaxation and all-atom force-field based normal mode analysis (NMA). We showed that combination of experimental and computational approach allows to identify flexible zones with higher level of confidence. We also found out that residues, that are involved VEGF-receptor interactions, reside in or close to the flexible zones, suggesting the critical role conformational plasticity plays in the non-covalent protein-protein interactions.
publishDate 2013
dc.date.none.fl_str_mv 2013
2013
2013
dc.type.none.fl_str_mv info:eu-repo/semantics/doctoralThesis
info:eu-repo/semantics/publishedVersion
format doctoralThesis
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/10803/113301
url http://hdl.handle.net/10803/113301
dc.language.none.fl_str_mv Inglés
language_invalid_str_mv Inglés
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv 163 p.
application/pdf
application/pdf
dc.publisher.none.fl_str_mv Universitat de Barcelona
publisher.none.fl_str_mv Universitat de Barcelona
dc.source.none.fl_str_mv TDX (Tesis Doctorals en Xarxa)
reponame:TDR. Tesis Doctorales en Red
instname:CBUC, CESCA
instname_str CBUC, CESCA
reponame_str TDR. Tesis Doctorales en Red
collection TDR. Tesis Doctorales en Red
repository.name.fl_str_mv
repository.mail.fl_str_mv
_version_ 1869414167181524992
spelling Molecular recognition in gas phase: theoretical and experimental study of non-covalent protein-ligand complexes by mass-spectrometryDyachenko, AndreyEspectrometria de massesEspectrometría de masasMass spectrometryInteraccions no covalentsInteracciones no covalentesNon-covalent interactionsInteraccions proteïna-proteïnaInteracciones proteína-proteínaProtein-protein interactionsInteraccions proteïna-lligandInteracciones proteína-ligandoLigand-protein interactionsCiències Experimentals i Matemàtiques547In the present thesis we have explored different factors that impede accurate quantitative description of non-covalent protein-protein and protein-ligand interactions and design of new potent and specific binders from the scratch. Firstly, we addressed the role of solvent in the mechanism of non-covalent interactions. Secondly, we tackled the question about the intrinsic conformational flexibility of the protein molecules and the part it plays in weak interactions between proteins. In the first part of the thesis we studied the interactions of vascular endothelial growth factor (VEGF) protein with five cyclic peptides in solution and gas phase. The results showed that affinities of five ligands to VEGF in solution and gas phase are ranked in inversed order. That is, the that has the highest affinity in solution (as shown by chemical shift perturbation NMR and isothermal titration calorimetry) forms the weakest complex with VEGF in gas phase, and vice versa. We compared gas-phase and solution binding affinities of of five peptides and made qualitative conclusions about the role of the solvent in protein-ligand interactions. In order to obtain more quantitative information about the gas-phase behavior of non-covalent complexes we have developed a combined experimental/theoretical approach to study the energetics of collisional activation of the ion prior to dissociation. We applied developed strategy to model CID in traveling wave ion guide (TWIG) collision cell. We validated the model on the CID of leu-enkephalin peptide and then applied developed strategy to five non-covalent protein-peptide complexes and found activation energies of their dissociation reactions. Next we applied ESI native MS to study the allosteric interactions between the molecular chaperonin GroEL and ATP. The obtained data allowed to construct a scheme of conformational transition of GroEL upon binding of ATP and distinguish between two different cooperativity models, providing strong arguments in favor of Monod-Wyman-Changeux (MWC) model. Finally, be studied the backbone dynamics of VEGF with a combination of NMR relaxation and all-atom force-field based normal mode analysis (NMA). We showed that combination of experimental and computational approach allows to identify flexible zones with higher level of confidence. We also found out that residues, that are involved VEGF-receptor interactions, reside in or close to the flexible zones, suggesting the critical role conformational plasticity plays in the non-covalent protein-protein interactions.Las biomoléculas de los organismos vivos realizan sus funciones principalmente a través de interacciones débiles reversibles entre ellas. La transducción de señal, la replicación de ADN/ARN, otros procesos enzimáticos y, virtualmente, cualquier otro proceso involucrado en las funciones vitales de cualquier organismo vivo (de las simples amebas, al complejo ser humano), requiere que las moléculas “hablen” entre ellas. Dicho lenguaje se basa en interacciones no covalentes. La flexibilidad conformacional es una propiedad esencial de las grandes biomoléculas, y muchas de las funciones desempeñadas por proteínas se basan en su capacidad para cambiar de conformación en respuesta a un factor externo. Geométricamente hablando, la presencia de flexibilidad en una proteína obstaculiza el diseño racional de medicamentos porque posibilita la existencia de un número muy elevado de conformaciones de dicha proteína. Por este motivo, cualquier información sobre la flexibilidad de una proteína es sumamente valiosa para la comprensión de PPI y PLI y para el diseño racional de medicamentos. Los capítulos 1-3 de la presente tesis versan sobre la solvatación, mientras que la flexibilidad se estudiara en el capitulo 4.Universitat de BarcelonaGiralt Lledó, ErnestUniversitat de Barcelona. Departament de Química Orgànica201320132013info:eu-repo/semantics/doctoralThesisinfo:eu-repo/semantics/publishedVersion163 p.application/pdfapplication/pdfhttp://hdl.handle.net/10803/113301TDX (Tesis Doctorals en Xarxa)reponame:TDR. Tesis Doctorales en Redinstname:CBUC, CESCAInglésADVERTIMENT. L'accés als continguts d'aquesta tesi doctoral i la seva utilització ha de respectar els drets de la persona autora. Pot ser utilitzada per a consulta o estudi personal, així com en activitats o materials d'investigació i docència en els termes establerts a l'art. 32 del Text Refós de la Llei de Propietat Intel·lectual (RDL 1/1996). Per altres utilitzacions es requereix l'autorització prèvia i expressa de la persona autora. En qualsevol cas, en la utilització dels seus continguts caldrà indicar de forma clara el nom i cognoms de la persona autora i el títol de la tesi doctoral. No s'autoritza la seva reproducció o altres formes d'explotació efectuades amb finalitats de lucre ni la seva comunicació pública des d'un lloc aliè al servei TDX. Tampoc s'autoritza la presentació del seu contingut en una finestra o marc aliè a TDX (framing). Aquesta reserva de drets afecta tant als continguts de la tesi com als seus resums i índexs.info:eu-repo/semantics/openAccessoai:www.tdx.cat:10803/1133012026-06-14T12:46:07Z
score 15,301603

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