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Modeling Kinetics of Protein-Ligand Systems
KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Theoretical Chemistry and Biology.ORCID iD: 0000-0003-4167-6413
2020 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Protein-ligand interactions dominate many life activities and are crucial for thedevelopment of tracers for diagnosing diseases and drugs for treating diseases.For protein-ligand interactions, the binding affinity is conventionally believedto be the most important indicator. However, there is increasing evidencethat the binding affinity alone is not sufficient for providing comprehensiveinformation about protein-ligand interactions. Kinetics, which describes theduration of the interactions and is closely related to the interaction mechanism,is considered as important as, or even more important than, the binding affinityin the study of the mechanisms of protein-ligand interactions.Although kinetics parameters of a protein-ligand system can be measuredexperimentally, the underlying molecular mechanism for the kinetics is difficultto reveal by experiment, which is, however, essential for understanding theorigin of the kinetics and for the rational design of drugs or tracers. In the lastdecade, computer simulations have emerged as a powerful tool for studying biomolecularsystems. Computer simulation methods have also been developedfor modeling kinetics of protein-ligand systems.In this thesis, I explored computer simulations for modeling kinetics propertiesof four different protein-ligand systems. In paper I, I studied the relationshipbetween the ligand binding and conformational changes of the ATAD2-BRD protein. In paper II, I investigated the free energy profile for the coupledfolding and binding of the intrinsically disordered protein p53 with MDM2and calculated the rate constants for the binding and unbinding processes. Inpaper III, I revealed the unbinding paths of the PET tracer ASEM from the  a7-nAChR, calculated the unbinding rate, and explored a way of how to findthe key protein conformational changes strongly coupled to the ligand unbindingprocess. In paper IV, I further refined our methodology for finding theunbinding paths and clarified the unbinding mechanism of the metabolite ofraloxifene from the enzyme CYP3A4.

Abstract [sv]

Protein-ligandinteraktioner dominerar många livsaktiviteter och är avgörande för utvecklingen av spårare för att diagnostisera sjukdomar och läkemedel för behandling av sjukdomar. För protein-ligandinteraktioner antas konventionell bindningsaffinitet vara den viktigaste indikatorn. Det finns emellertid ökande bevis på att bindningsaffiniteten enbart inte är tillräcklig för att tillhandahålla omfattande information om protein-ligandinteraktioner. Kinetik, som beskriver varaktigheten på interaktioner och är nära besläktad med interaktionsmekanismen, anses vara så viktig som, eller ännu viktigare än bindningsaffiniteten i studien av mekanismerna för protein-ligandinteraktioner.

 

Även om kinetikparametrar i ett protein-ligandsystem kan mätas experimentellt är den underliggande molekylära mekanismen för kinetiken svår att avslöja genom experiment, vilket dock är väsentligt för att förstå kinetikens ursprung och för den rationella utformningen av läkemedel eller spårare . Under det senaste decenniet har datorsimuleringar framkommit som ett kraftfullt verktyg för att studera biomolekylära system. Datorsimuleringsmetoder har också utvecklats för att modellera kinetik för protein-ligandsystem.

 

I den här avhandlingen undersökte jag datorsimuleringar för modellering av kinetiska egenskaper hos fyra olika protein-ligandsystem. I papper I studerade jag sambandet mellan ligandbindningen och konformationella förändringar av ATAD2-BRD-proteinet. I papper II undersökte jag den fria energiprofilen för den kopplade vikningen och bindningen av det intrinsiskt störda proteinet p53-peptid med MDM2 och beräknade hastighetskonstanterna för bindnings- och bindningsförfarandena. I papper III avslöjade jag de bindande vägarna för PET-spåraren ASEM från α7-nAChR, beräknade bindningsgraden och utforskade ett sätt att hitta de viktiga proteinkonformationella förändringarna starkt kopplade till ligandbindningsprocessen. I papper IV förfinade jag ytterligare vår metod för att hitta de bindande vägarna och klargjorde den bindande mekanismen för metaboliten av raloxifen från enzymet CYP3A4.

Place, publisher, year, edition, pages
KTH Royal Institute of Technology, 2020. , p. 57
Series
TRITA-CBH-FOU ; 2020:25
National Category
Theoretical Chemistry
Research subject
Theoretical Chemistry and Biology
Identifiers
URN: urn:nbn:se:kth:diva-273146ISBN: 978-91-7873-545-7 (print)OAI: oai:DiVA.org:kth-273146DiVA, id: diva2:1429036
Public defence
2020-06-03, https://kth-se.zoom.us/webinar/register/WN_ZxwH8-GQTFaTv9ifiiTsAA ​, Stockholm, 10:00 (English)
Opponent
Supervisors
Note

QC 2020-05-08

Available from: 2020-05-08 Created: 2020-05-07 Last updated: 2020-05-20Bibliographically approved
List of papers
1. Mechanistic insights into peptide and ligand binding of the ATAD2-bromodomain via atomistic simulations disclosing a role of induced fit and conformational selection
Open this publication in new window or tab >>Mechanistic insights into peptide and ligand binding of the ATAD2-bromodomain via atomistic simulations disclosing a role of induced fit and conformational selection
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2018 (English)In: Physical Chemistry, Chemical Physics - PCCP, ISSN 1463-9076, E-ISSN 1463-9084, Vol. 20, no 36, p. 23222-23232Article in journal (Refereed) Published
Abstract [en]

ATAD2 has emerged as a promising bromodomain (BRD)-containing therapeutic drug target in multiple human cancers. However, recent druggability assessment studies predicted ATAD2's BRD as a target 'difficult to drug' because its binding pocket possesses structural features that are unfeasible for ligand binding. Here, by using all-atom molecular dynamics simulations and an advanced metadynamics method, we demonstrate a dynamic view of the binding pocket features which can hardly be obtained from the "static" crystal data. The most important features disclosed from our simulation data, include: (1) a distinct 'open-to-closed' conformational switch of the ZA loop region in the context of peptide or ligand binding, akin to the induced fit mechanism of molecular recognition, (2) a dynamic equilibrium of the BC loop "in" and "out" conformations, highlighting a role in the conformational selection mechanism for ligand binding, and (3) a new binding region identified distal to the histone-binding pocket that might have implications in bromodomain biology and in inhibitor development. Moreover, based on our simulation results, we propose a model for an "auto-regulatory" mechanism of ATAD2's BRD for histone binding. Overall, the results of this study will not only have implications in bromodomain biology but also provide a theoretical basis for the discovery of new ATAD2's BRD inhibitors.

Place, publisher, year, edition, pages
Royal Society of Chemistry, 2018
Keywords
AAA protein, ATAD2 protein, human, DNA binding protein, ligand, peptide, binding site, chemistry, conformation, human, molecular dynamics, ATPases Associated with Diverse Cellular Activities, Binding Sites, DNA-Binding Proteins, Humans, Ligands, Molecular Conformation, Molecular Dynamics Simulation, Peptides
National Category
Atom and Molecular Physics and Optics
Identifiers
urn:nbn:se:kth:diva-236425 (URN)10.1039/c8cp03860k (DOI)000447370600005 ()30137066 (PubMedID)2-s2.0-85053795262 (Scopus ID)
Note

QC 20181026

Available from: 2018-10-26 Created: 2018-10-26 Last updated: 2020-05-07Bibliographically approved
2. Free Energy Profile and Kinetics for Coupled Folding and Binding of the Intrinsically Disordered Protein p53 with MDM2
Open this publication in new window or tab >>Free Energy Profile and Kinetics for Coupled Folding and Binding of the Intrinsically Disordered Protein p53 with MDM2
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

Intrinsically disordered proteins (IDPs) exert their functions by binding to partner proteins via a complex process that includes coupled folding and binding. Motivated by that inhibiting the binding of the IDP p53 to its partner MDM2 has become a promising strategy for drug design and that understanding of this process poses a most significant challenging task, we present an atomistic level simulation of the coupled folding and binding process linking the IDP p53 to MDM2. Using bias-exchange metadynamics (BE-MetaD) and infrequent metadynamics (InMetaD) we estimate the binding free energy, the unbinding rate and the binding rate. By analyzing the stable intermediates, we uncover the role of nonnative interactions played in the p53-MDM2 binding/unbinding process. We use a three-state model to describe the whole binding/unbinding process and to obtain the corresponding rate constants. Our work shows that the binding of p53 favors an induced fit mechanism which proceeds in a stepwise fashion. In general, InMetaD gave consistent results with BE-MetaD in terms of binding mechanism and intermediates, proving the robustness of our studies of the p53-MDM2 system using metadynamics. The results contribute to the in-depth understanding for the coupled folding and binding process that is needed for the design of MDM2 inhibitors.

Keywords
p53-MDM2, free energy landscape, kinetics, bias-exchange metadynamics, infrequent metadynamics
National Category
Natural Sciences
Identifiers
urn:nbn:se:kth:diva-251311 (URN)
Note

QC 20190619

Available from: 2019-05-10 Created: 2019-05-10 Last updated: 2020-05-07Bibliographically approved
3. Enhanced Sampling Simulations of Ligand Unbinding Kinetics Controlled by Protein Conformational Changes
Open this publication in new window or tab >>Enhanced Sampling Simulations of Ligand Unbinding Kinetics Controlled by Protein Conformational Changes
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2019 (English)In: Journal of Chemical Information and Modeling, ISSN 1549-9596, E-ISSN 1549-960X, Vol. 59, no 9, p. 3910-3918Article in journal (Refereed) Published
Abstract [en]

Understanding unbinding kinetics of protein-ligand systems is of great importance for the design of ligands with desired specificity and safety. In recent years, enhanced sampling techniques have emerged as effective tools for studying unbinding kinetics of protein-ligand systems at the atomistic level. However, in many protein-ligand systems, the ligand unbinding processes are strongly coupled to protein conformational changes and the disclosure of the hidden degrees of freedom closely related to the protein conformational changes so that sampling is enhanced over these degrees of freedom remains a great challenge. Here, we show how potential-scaled molecular dynamics (sMD) and infrequent metadynamics (InMetaD) simulation techniques can be combined to successfully reveal the unbinding mechanism of 3-(1,4-diazabicyclo[3.2.2]nonan-4-yl)-6-[F-18]fluorodibenzo[b,d]thiophen e 5,5-dioxide ([F-18]ASEM) from a chimera structure of the alpha 7-nicotinic acetylcholine receptor. By using sMD simulations, we disclosed that the "close to "open" conformational change of loop C plays a key role in the ASEM unbinding process. By carrying out InMetaD simulations with this conformational change taken into account as an additional collective variable, we further captured the key states in the unbinding process and clarified the unbinding mechanism of ASEM from the protein. Our work indicates that combining sMD and InMetaD simulation techniques can be an effective approach for revealing the unbinding mechanism of a protein-ligand system where protein conformational changes control the unbinding process.

Place, publisher, year, edition, pages
AMER CHEMICAL SOC, 2019
National Category
Chemical Sciences
Identifiers
urn:nbn:se:kth:diva-262805 (URN)10.1021/acs.jcim.9b00523 (DOI)000487769800034 ()31454236 (PubMedID)2-s2.0-85072587758 (Scopus ID)
Note

QC 20191021

Available from: 2019-10-21 Created: 2019-10-21 Last updated: 2020-05-07Bibliographically approved
4. An integrated enhanced sampling approach to characterize the unbinding process of diquinone methide from cytochrome P450 3A4
Open this publication in new window or tab >>An integrated enhanced sampling approach to characterize the unbinding process of diquinone methide from cytochrome P450 3A4
(English)Manuscript (preprint) (Other academic)
Abstract [en]

The catalytic properties of an enzyme are determined not only by the mechanism of the reaction taking place at the active center, but also by how fast the substrate enters and the product releases from the enzyme. Revealing the mechanism of a substrate unbinding from an enzyme is therefore of great significance for guiding enzyme engineering or for the rational design of drug molecules. When the active site is deeply buried in an enzyme and the protein conformational changes play a crucial role in regulating the unbinding kinetics, understanding the mechanism of the unbinding process presents a great challenge both theoretically and experimentally. Here we show how to combine classic molecular dynamic simulations, enhanced sampling approaches, free energy calculations, and quantum chemistry calculations to clarify the mechanisms of diquinone methide (DQR), the metabolite product of drug Raloxifene, unbinding from Cytochrome P450 (CYP) 3A4 and the suicide inactivation of CYP3A4 by raloxifene. Our studies indicate that DQR may have more than one unbinding pathways from the active site of CYP3A4. We found that several intermediate states, which are strongly coupled to the conformational changes of CYP3A4, were involved in the DQR unbinding process. In one of the intermediate states, DQR can covalently bind to Cys239 of CYP3A4, which hinders the substrate from entering or leaving the enzyme. Our work thus reveals the mechanisms of DQR unbinding from CYP3A4 and the suicide inactivation of CYP3A4 by raloxifene.

National Category
Theoretical Chemistry
Identifiers
urn:nbn:se:kth:diva-273155 (URN)
Note

QC 20200511

Available from: 2020-05-07 Created: 2020-05-07 Last updated: 2020-05-11Bibliographically approved

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