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Ultrafast Vibrational Spectroscopy of Allosteric Proteins

English title Ultrafast Vibrational Spectroscopy of Allosteric Proteins
Applicant Hamm Peter
Number 165789
Funding scheme Project funding
Research institution Institut für Chemie Universität Zürich
Institution of higher education University of Zurich - ZH
Main discipline Physical Chemistry
Start/End 01.01.2017 - 31.12.2019
Approved amount 773'577.00
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Keywords (5)

Infrared Spectroscopy; 2D IR spectroscopy; Protein Dynamics; Allostery; Ultrafast Spectrosocpy

Lay Summary (German)

Lead
Allosterie ist ein wichtiges Konzept in der Biochemie, das für die Regulation chemischer Prozesse in einer Zelle verantwortlich ist. Allosterische Proteine besitzen in der Regel zwei Bindungszentren; ein enzymatisches Zentrum, in dem ein bestimmtes Molekül chemisch modifiziert wird, sowie ein allosterisches Zentrum. In Abhängigkeit davon, ob ein bestimmtes anderes Molekül (Ligand) an dieses allosterische Zentrum bindet, wird die chemische Aktivität im enzymatischen Zentrum hoch- bzw. runtergefahren. Oft sind die beiden Bindungszentren relativ weit voneinander entfernt, und bis heute ist weitgehend unklar, wie sie miteinander „kommunizieren“. Man geht davon aus, dass ein Ligand im allosterischen Zentrum eine, wenn auch oft sehr kleine, strukturelle Veränderung des Proteins als Ganzes verursacht, die einen Einfluss auf die chemische Aktivität des enzymatischen Zentrums hat.
Lay summary

Lead

Allosterie  ist ein wichtiges Konzept in der Biochemie, das für die Regulation chemischer Prozesse in einer Zelle verantwortlich ist. Allosterische Proteine besitzen in der Regel zwei Bindungszentren; ein enzymatisches Zentrum, in dem ein bestimmtes Molekül chemisch modifiziert wird, sowie ein allosterisches Zentrum. In Abhängigkeit davon, ob ein bestimmtes anderes Molekül (Ligand) an dieses allosterische Zentrum bindet, wird die chemische Aktivität im enzymatischen Zentrum hoch- bzw. runtergefahren. Oft sind die beiden  Bindungszentren relativ weit voneinander entfernt, und bis heute ist weitgehend unklar, wie sie miteinander „kommunizieren“. Man geht davon aus, dass ein Ligand im allosterischen Zentrum eine, wenn auch oft sehr kleine, strukturelle Veränderung des Proteins als Ganzes verursacht, die einen Einfluss auf die chemische Aktivität des enzymatischen Zentrums hat.

 

Inhalt und Ziel des Forschungsprojekts

Wir entwickeln speziell designte Proteine und/oder Liganden, um mit Hilfe der zeitaufgelösten Infrarotspektroskopie die Ausbreitung eines Signals in einem Protein messen zu können. Dazu stören wir das Protein gezielt im Bereich des allosterischen Zentrums in dem wir z.B. einen durch einen kurzen Lichtblitz schaltbaren Liganden vom Protein dissoziieren, und messen die spektroskopische Antwort einer künstlichen Aminosäure mit einer sehr charakteristische Infrarotabsorptionsbande in Abhängigkeit vom Abstand vom allosterischen Zentrum.

 

Wissenschaftlicher und gesellschaftlicher Kontext des Forschungsprojekts

Sehr viele Medikamente wirken, indem sie die allosterische Regulation bestimmter Proteine beeinflussen; z.B. indem sie das allosterisches Zentrum blockieren. Ein tieferes Verständnis des Mechanismus von Allosterie wird es erlauben, bessere Medikamente zu designen, anstelle, wie heute meist üblich, grosse Moleküldatenbanken zu screenen.    

Direct link to Lay Summary Last update: 21.07.2016

Employees

Publications

Publication
Using azobenzene photocontrol to set proteins in motion
BozovicOlga, JankovicBrankica, HammPeter (2021), Using azobenzene photocontrol to set proteins in motion, in NATURE REVIEWS CHEMISTRY, 6(2), 112-124.
Photocontrolling Protein–Peptide Interactions: From Minimal Perturbation to Complete Unbinding
Jankovic Brankica, Gulzar Adnan, Zanobini Claudio, Bozovic Olga, Wolf Steffen, Stock Gerhard, Hamm Peter (2019), Photocontrolling Protein–Peptide Interactions: From Minimal Perturbation to Complete Unbinding, in Journal of the American Chemical Society, 141(27), 10702-10710.
Azidohomoalanine: A Minimally Invasive, Versatile, and Sensitive Infrared Label in Proteins To Study Ligand Binding
Zanobini Claudio, Bozovic Olga, Jankovic Brankica, Koziol Klemens L., Johnson Philip J. M., Hamm Peter, Gulzar Adnan, Wolf Steffen, Stock Gerhard (2018), Azidohomoalanine: A Minimally Invasive, Versatile, and Sensitive Infrared Label in Proteins To Study Ligand Binding, in The Journal of Physical Chemistry B, 122(44), 10118-10125.
Aqueous solvation from the water perspective
Ahmed Saima, Pasti Andrea, Fernández-Terán Ricardo J., Ciardi Gustavo, Shalit Andrey, Hamm Peter (2018), Aqueous solvation from the water perspective, in The Journal of Chemical Physics, 148(23), 234505-234505.
A non-equilibrium approach to allosteric communication
Stock Gerhard, Hamm Peter (2018), A non-equilibrium approach to allosteric communication, in Philosophical Transactions of the Royal Society B: Biological Sciences, 373(1749), 20170187-20170187.
2D-IR Spectroscopy of an AHA Labeled Photoswitchable PDZ2 Domain
Stucki-Buchli Brigitte, Johnson Philip J. M., Bozovic Olga, Zanobini Claudio, Koziol Klemens L., Hamm Peter, Gulzar Adnan, Wolf Steffen, Buchenberg Sebastian, Stock Gerhard (2017), 2D-IR Spectroscopy of an AHA Labeled Photoswitchable PDZ2 Domain, in The Journal of Physical Chemistry A, 121(49), 9435-9445.
Quantifying Biomolecular Recognition with Site-Specific 2D Infrared Probes
Johnson Philip J. M., Koziol Klemens L., Hamm Peter (2017), Quantifying Biomolecular Recognition with Site-Specific 2D Infrared Probes, in The Journal of Physical Chemistry Letters, 8(10), 2280-2284.
Solvation Layer of Antifreeze Proteins Analyzed with a Markov State Model,
Wellig Sebastian, Hamm Peter, Solvation Layer of Antifreeze Proteins Analyzed with a Markov State Model,, in J. Phys. Chem. B.

Collaboration

Group / person Country
Types of collaboration
Prof. Gerhard Stock, University of Freiburg Germany (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
Prof. Amedeo Caflisch, University of Zurich Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication

Scientific events

Active participation

Title Type of contribution Title of article or contribution Date Place Persons involved
Seminar, University of Pennsylvania Individual talk A Nonequilibrium Approach to Allosteric Communication 24.10.2019 Philadelphia, United States of America Hamm Peter;
TRVS2019-Time resolved vibrational spectroscopy Talk given at a conference A Non-equilibrium Approach to Allosteric Communication 08.09.2019 Auckland, New Zealand Bozovic Olga;
SCS Fall Meeting 2019 Poster Transient infrared response of a PDZ2 domain protein upon light induced ligand perturbation 06.09.2019 Zürich, Switzerland Bozovic Olga; Zanobini Claudio;
SCS Fall Meeting Poster Light-induced peptide unbinding in the RNase S complex 06.09.2019 Zürich, Switzerland Jankovic Brankica;
SCS Fall Meeting Poster Transient infrared response of a PDZ2 domain protein upon light induced ligand perturbation 06.09.2019 Zürich, Switzerland Jankovic Brankica;
ESULaB 2019 Talk given at a conference A Nonequilibrium Approach to Allosteric Communication 04.09.2019 Jena, Germany Hamm Peter;
Colloquium, SFB 1078, Freie Universität Berlin,Germany Individual talk A Nonequilibrium Approach to Allosteric Communication 20.05.2019 Berlin, Germany Hamm Peter;
MUST Annual Meeting Poster Transient infrared response of a PDZ2 domain protein upon light induced ligand perturbation, 14.01.2019 Grindelwald, Switzerland Bozovic Olga; Zanobini Claudio;
9th NCCR MUST Annual Meeting Poster Transient infrared response of PDZ2 domain protein upon light induced ligand perturbation 14.01.2019 Grindelwald, Switzerland Jankovic Brankica;
MUST Annual Meeting Poster Distance dependence of vibrational energy transfer: calibrating a molecular ruler with 2D ATR-IR spectroscopy 14.01.2019 Grindelwald, Switzerland Bozovic Olga;
9th NCCR MUST Annual Meeting Poster Investigating S-protein/S-peptide structural changes in time-resolved manner 14.01.2019 Grindelwald, Switzerland Jankovic Brankica;
Swiss Chemical Society Fall Meeting Poster Photo-controlling protein allostery in PDZ3 domain 07.09.2018 Lausanne, Switzerland Jankovic Brankica;
Swiss Chemical Society Fall Meeting Poster Modulating S-protein/S-peptide interactions in RNase S complex by light 07.09.2018 Lausanne, Switzerland Bozovic Olga; Zanobini Claudio;
Swiss Chemical Society Fall Meeting Poster Modulating S-protein/S-peptide interactions in RNase S complex by light 07.09.2018 Lausanne, Switzerland Jankovic Brankica;
Swiss Chemical Society Fall Meeting Poster , Photo-controlling protein allostery in PDZ3 domain 07.09.2018 Lausanne, Switzerland Zanobini Claudio; Bozovic Olga;
8th Annual Meeting of the NCCR MUST Poster Light induced conformational perturbation of S-protein/S-peptide complex 22.01.2018 Grindelwald, Switzerland Zanobini Claudio; Bozovic Olga;
MUST Annual Meeting Poster Light induced conformational perturbation of S-protein/S-peptide complex 22.01.2018 Grindelwald, Switzerland Jankovic Brankica;
MUST Annual Meeting Poster Photo-controlling protein allostery in PDZ3 domain 22.01.2018 Grindelwald, Switzerland Jankovic Brankica;
8th Annual Meeting of the NCCR MUST Poster Photo-controlling protein allostery in PDZ3 domain 22.01.2018 Grindelwald, Switzerland Bozovic Olga; Zanobini Claudio;
NCCR MUST annual meeting Poster photo-controling protein allostery in PDZ3 domain 22.01.2018 Grindelwald, Switzerland Bozovic Olga;
SCS fall meeting Poster Investigation of PDZ2/RagEF peptide interactions by 2D infrared spectroscopy using wild type and modified photoswitchable peptide ligands 21.08.2017 Bern, Switzerland Bozovic Olga;
Swiss Chemical Society Fall Meeting Poster Investigation of PDZ2/RA-GEF2 peptide interactions by 2DIR spectroscopy using wild type and modified photoswitchable peptide ligands 21.08.2017 Bern, Switzerland Jankovic Brankica;
Swiss Chemical Society Fall Meeting Poster Light induced conformational isomerisation of helical photoswitchable S-peptide and perturbation of S-protein/S-peptide complex 21.08.2017 Bern, Switzerland Jankovic Brankica;
Time Resolved Vibrational Spectroscopy Meeting Poster Investigation of peptide interactions by 2DIR spectroscopy using wild type and modified photoswitchable peptide ligands 16.07.2017 Cambridge, Great Britain and Northern Ireland Jankovic Brankica;
TRVS Cambridge Poster "Investigation of PDZ2/RagEF peptide interactions by 2D infrared spectroscopy using wild type and modified photoswitchable peptide ligands 16.07.2017 Cambridge, Great Britain and Northern Ireland Bozovic Olga;
Doktorandentag 2017 Poster Investigating protein-ligand interactions using two-dimensional infrared spectroscopy with photoswitchable peptide ligand 29.06.2017 Zürich, Switzerland Zanobini Claudio;
Doktorandentag 2017 Poster Two-dimensional infrared spectroscopy of a site-specifically labeled photoswitchable allosteric protein 29.06.2017 Zürich, Switzerland Bozovic Olga;
MUST Annual Meeting 2017 Poster , Investigating protein-ligand interactions using two-dimensional infrared spectroscopy with photoswitchable peptide ligand 08.01.2017 Grindelwald, Switzerland Zanobini Claudio;


Associated projects

Number Title Start Funding scheme
188694 Ultrafast Vibrational Spectroscopy of Allosteric Proteins (Extension) 01.01.2020 Project funding

Abstract

Allostery is the coupling of two separated sites of a protein, where binding of a ligand at the so-called allosteric site changes the function of the protein at a remote active site. The very question how these two sites communicate with each other is largely unanswered and is an important topic that is heavily studied from many perspectives. Transient THz, IR and 2D IR spectroscopy provides the necessary time resolution and chemical selectivity to follow the propagation of a signal after an external perturbation, which we will induce by an ultrashort laser pulse at the allosteric site of an allosteric protein. In detail, we will use three complementary approaches:- Building on results from an expired ERC Advanced grant, we will vibrationally label a photo-switchable PDZ2 domain at various positions throughout the protein. In this case, the perturbation is induced via a photo-switch (i.e., a azobenzene-derivative), which cross-links the binding groove of that allosteric protein in a way that the \textit{cis-trans} isomerisation of the photo-switch induces a conformational change of the protein that mimics ligand binding. Transient IR and 2D IR spectroscopy of the vibrational labels will allow us to follow the pathway of the signal propagating through the protein.- The more direct way of measuring the protein response after ligand binding/unbinding requires a ligand whose binding affinity can be switched, i.e., with the photo-switch attached to the ligand. To that end, we will design a ligand that has a high binding affinity in the one state of the photo-switch and smaller binding affinity in the other state, so that activating the photo-switch will initiate ligand unbinding. The protein will again be vibrationally labeled and measured by transient IR and 2D IR spectroscopy.- By transient THz spectroscopy, we will experimentally test a previous hypothesis originating from molecular dynamics simulations, according to which the water solvation shell rearranges significantly upon switching the PDZ2 domain, despite the fact that the structural change of the protein is relatively small. We proposed that this rearrangement of the solvation layer is a possible mechanism of allostery. In this experiment, we will probe selectively the change of the hydrogen-bond network of waters in the solvation layer of the protein.We are convinced that these experiments, complemented with molecular dynamics simulations, will provide an unprecedented new view into the mechanism of allostery, and as such, eventually, pave the way for a more rational design of new drugs that interact with allosteric regulation networks.
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