Bachelor of Science, University Of Helsinki (2013)
Master of Science, University Of Helsinki (2014)
Doctor of Philosophy, University Of Helsinki (2018)
How GPCR Phosphorylation Patterns Orchestrate Arrestin-Mediated Signaling.
Binding of arrestin to phosphorylated G-protein-coupled receptors (GPCRs) controls many aspects of cell signaling. The number and arrangement of phosphates may vary substantially for a given GPCR, and different phosphorylation patterns trigger different arrestin-mediated effects. Here, we determine how GPCR phosphorylation influences arrestin behavior by using atomic-level simulations and site-directed spectroscopy to reveal the effects of phosphorylation patterns on arrestin binding and conformation. We find that patterns favoring binding differ from those favoring activation-associated conformational change. Both binding and conformation depend more on arrangement of phosphates than on their total number, with phosphorylation at different positions sometimes exerting opposite effects. Phosphorylation patterns selectively favor a wide variety of arrestin conformations, differently affecting arrestin sites implicated in scaffolding distinct signaling proteins. We also reveal molecular mechanisms of these phenomena. Our work reveals the structural basis for the long-standing "barcode" hypothesis and has important implications for design of functionally selective GPCR-targeted drugs.
View details for DOI 10.1016/j.cell.2020.11.014
View details for PubMedID 33296703
Angiotensin and biased analogs induce structurally distinct active conformations within a GPCR.
Science (New York, N.Y.)
2020; 367 (6480): 888–92
Biased agonists of G protein-coupled receptors (GPCRs) preferentially activate a subset of downstream signaling pathways. In this work, we present crystal structures of angiotensin II type 1 receptor (AT1R) (2.7 to 2.9 angstroms) bound to three ligands with divergent bias profiles: the balanced endogenous agonist angiotensin II (AngII) and two strongly beta-arrestin-biased analogs. Compared with other ligands, AngII promotes more-substantial rearrangements not only at the bottom of the ligand-binding pocket but also in a key polar network in the receptor core, which forms a sodium-binding site in most GPCRs. Divergences from the family consensus in this region, which appears to act as a biased signaling switch, may predispose the AT1R and certain other GPCRs (such as chemokine receptors) to adopt conformations that are capable of activating beta-arrestin but not heterotrimeric Gq protein signaling.
View details for DOI 10.1126/science.aay9813
View details for PubMedID 32079768
Molecular mechanism of biased signaling in a prototypical G protein-coupled receptor.
Science (New York, N.Y.)
2020; 367 (6480): 881–87
Biased signaling, in which different ligands that bind to the same G protein-coupled receptor preferentially trigger distinct signaling pathways, holds great promise for the design of safer and more effective drugs. Its structural mechanism remains unclear, however, hampering efforts to design drugs with desired signaling profiles. Here, we use extensive atomic-level molecular dynamics simulations to determine how arrestin bias and G protein bias arise at the angiotensin II type 1 receptor. The receptor adopts two major signaling conformations, one of which couples almost exclusively to arrestin, whereas the other also couples effectively to a G protein. A long-range allosteric network allows ligands in the extracellular binding pocket to favor either of the two intracellular conformations. Guided by this computationally determined mechanism, we designed ligands with desired signaling profiles.
View details for DOI 10.1126/science.aaz0326
View details for PubMedID 32079767
Conformational transitions of a neurotensin receptor1-Gi1complex.
Neurotensin receptor1 (NTSR1) is a G-protein-coupled receptor (GPCR) that engages multiple subtypes of G protein, and is involved in the regulation of blood pressure, body temperature, weight and the response to pain. Here we present structures of human NTSR1 in complex with the agonist JMV449 and the heterotrimeric Gi1 protein, at a resolution of 3A. We identify two conformations: a canonical-state complex that is similar to recently reported GPCR-Gi/o complexes (in which the nucleotide-binding pocket adopts moreflexible conformations that may facilitate nucleotide exchange), and a non-canonical state in which the G protein is rotated by about 45degrees relative to the receptor and exhibits a more rigid nucleotide-binding pocket. In the non-canonical state, NTSR1 exhibits features of both active and inactive conformations, which suggests that the structure may represent an intermediate form along the activation pathway of G proteins. This structural information, complemented by molecular dynamics simulations and functional studies, provides insights into the complex process of G-protein activation.
View details for DOI 10.1038/s41586-019-1337-6
View details for PubMedID 31243364
Energetics and dynamics of a light-driven sodium-pumping rhodopsin
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
2017; 114 (27): 7043–48
The conversion of light energy into ion gradients across biological membranes is one of the most fundamental reactions in primary biological energy transduction. Recently, the structure of the first light-activated Na+ pump, Krokinobacter eikastus rhodopsin 2 (KR2), was resolved at atomic resolution [Kato HE, et al. (2015) Nature 521:48-53]. To elucidate its molecular mechanism for Na+ pumping, we perform here extensive classical and quantum molecular dynamics (MD) simulations of transient photocycle states. Our simulations show how the dynamics of key residues regulate water and ion access between the bulk and the buried light-triggered retinal site. We identify putative Na+ binding sites and show how protonation and conformational changes gate the ion through these sites toward the extracellular side. We further show by correlated ab initio quantum chemical calculations that the obtained putative photocycle intermediates are in close agreement with experimental transient optical spectroscopic data. The combined results of the ion translocation and gating mechanisms in KR2 may provide a basis for the rational design of novel light-driven ion pumps with optogenetic applications.
View details for DOI 10.1073/pnas.1703625114
View details for Web of Science ID 000404576100060
View details for PubMedID 28611220
View details for PubMedCentralID PMC5502629
Autoantibody mimicry of hormone action at the thyrotropin receptor.
Thyroid hormones are vital to metabolism, growth and development1. Thyroid hormone synthesis is controlled by thyrotropin (TSH), which acts at the thyrotropin receptor (TSHR)2. Autoantibodies that activate the TSHR pathologically increase thyroid hormones in Graves' disease3. How autoantibodies mimic TSH function remains unclear. We determined cryogenic-electron microscopy structures of active and inactive TSHR. In inactive TSHR, the extracellular domain lies close to the membrane bilayer. TSH selects an upright orientation of the extracellular domain due to steric clashes between a conserved hormone glycan and the membrane bilayer. An activating autoantibody from a Graves' disease patient selects a similar upright orientation of the extracellular domain. Reorientation of the extracellular domain transduces a conformational change in the seven transmembrane domain via a conserved hinge domain, a tethered peptide agonist, and a phospholipid that binds within the seven transmembrane domain. Rotation of the TSHR extracellular domain relative to the membrane bilayer is sufficient for receptor activation, revealing a shared mechanism for other glycoprotein hormone receptors that may also extend to other G protein-coupled receptors with large extracellular domains.
View details for DOI 10.1038/s41586-022-05159-1
View details for PubMedID 35940205
Cryo-EM, Protein Engineering, and Simulation Enable the Development of Peptide Therapeutics against Acute Myeloid Leukemia.
ACS central science
2022; 8 (2): 214-222
Cryogenic electron microscopy (cryo-EM) has emerged as a viable structural tool for molecular therapeutics development against human diseases. However, it remains a challenge to determine structures of proteins that are flexible and smaller than 30 kDa. The 11 kDa KIX domain of CREB-binding protein (CBP), a potential therapeutic target for acute myeloid leukemia and other cancers, is a protein which has defied structure-based inhibitor design. Here, we develop an experimental approach to overcome the size limitation by engineering a protein double-shell to sandwich the KIX domain between apoferritin as the inner shell and maltose-binding protein as the outer shell. To assist homogeneous orientations of the target, disulfide bonds are introduced at the target-apoferritin interface, resulting in a cryo-EM structure at 2.6 A resolution. We used molecular dynamics simulations to design peptides that block the interaction of the KIX domain of CBP with the intrinsically disordered pKID domain of CREB. The double-shell design allows for fluorescence polarization assays confirming the binding between the KIX domain in the double-shell and these interacting peptides. Further cryo-EM analysis reveals a helix-helix interaction between a single KIX helix and the best peptide, providing a possible strategy for developments of next-generation inhibitors.
View details for DOI 10.1021/acscentsci.1c01090
View details for PubMedID 35233453
Atypical structural snapshots of human cytomegalovirus GPCR interactions with host G proteins.
1800; 8 (3): eabl5442
Human cytomegalovirus (HCMV) encodes G protein-coupled receptors (GPCRs) US28 and US27, which facilitate viral pathogenesis through engagement of host G proteins. Here we report cryo-electron microscopy structures of US28 and US27 forming nonproductive and productive complexes with Gi and Gq, respectively, exhibiting unusual features with functional implications. The "orphan" GPCR US27 lacks a ligand-binding pocket and has captured a guanosine diphosphate-bound inactive Gi through a tenuous interaction. The docking modes of CX3CL1-US28 and US27 to Gi favor localization to endosome-like curved membranes, where US28 and US27 can function as nonproductive Gi sinks to attenuate host chemokine-dependent Gi signaling. The CX3CL1-US28-Gq/11 complex likely represents a trapped intermediate during productive signaling, providing a view of a transition state in GPCR-G protein coupling for signaling. Our collective results shed new insight into unique G protein-mediated HCMV GPCR structural mechanisms, compared to mammalian GPCR counterparts, for subversion of host immunity.
View details for DOI 10.1126/sciadv.abl5442
View details for PubMedID 35061538
Selective G protein signaling driven by substance P-neurokinin receptor dynamics.
Nature chemical biology
The neuropeptide substance P (SP) is important in pain and inflammation. SP activates the neurokinin-1 receptor (NK1R) to signal via Gq and Gs proteins. Neurokinin A also activates NK1R, but leads to selective Gq signaling. How two stimuli yield distinct G protein signaling at the same G protein-coupled receptor remains unclear. We determined cryogenic-electron microscopy structures of active NK1R bound to SP or the Gq-biased peptide SP6-11. Peptide interactions deep within NK1R are critical for receptor activation. Conversely, interactions between SP and NK1R extracellular loops are required for potent Gs signaling but not Gq signaling. Molecular dynamics simulations showed that these superficial contacts restrict SP flexibility. SP6-11, which lacks these interactions, is dynamic while bound to NK1R. Structural dynamics of NK1R agonists therefore depend on interactions with the receptor extracellular loops and regulate G protein signaling selectivity. Similar interactions between other neuropeptides and their cognate receptors may tune intracellular signaling.
View details for DOI 10.1038/s41589-021-00890-8
View details for PubMedID 34711980
- Absorption shifts of diastereotopically ligated chlorophyll dimers of photosystem I PHYSICAL CHEMISTRY CHEMICAL PHYSICS 2019; 21 (13): 6851–58
Tuning the Protein-Induced Absorption Shifts of Retinal in Engineered Rhodopsin Mimics
CHEMISTRY-A EUROPEAN JOURNAL
2016; 22 (24): 8254–61
Rational design of light-capturing properties requires understanding the molecular and electronic structure of chromophores in their native chemical or biological environment. We employ here large-scale quantum chemical calculations to study the light-capturing properties of retinal in recently designed human cellular retinol binding protein II (hCRBPII) variants (Wang et al. Science, 2012, 338, 1340-1343). Our calculations show that these proteins absorb across a large part of the visible spectrum by combined polarization and electrostatic effects. These effects stabilize the ground or excited state energy levels of the retinal by perturbing the Schiff-base or β-ionone moieties of the chromophore, which in turn modulates the amount of charge transfer within the molecule. Based on the predicted tuning principles, we design putative in silico mutations that further shift the absorption properties of retinal in hCRBPII towards the ultraviolet and infrared regions of the spectrum.
View details for DOI 10.1002/chem.201505126
View details for Web of Science ID 000380269400029
View details for PubMedID 27120137
Exploring the Light-Capturing Properties of Photosynthetic Chlorophyll Clusters Using Large-Scale Correlated Calculations
JOURNAL OF CHEMICAL THEORY AND COMPUTATION
2016; 12 (6): 2644–51
Chlorophylls are light-capturing units found in photosynthetic proteins. We study here the ground and excited state properties of monomeric, dimeric, and tetrameric models of the special chlorophyll/bacteriochlorophyll (Chl/BChl) pigment (P) centers P700 and P680/P870 of type I and type II photosystems, respectively. In the excited state calculations, we study the performance of the algebraic diagrammatic construction through second-order (ADC(2)) method in combination with the reduced virtual space (RVS) approach and the recently developed Laplace-transformed scaled-opposite-spin (LT-SOS) algorithm, which allows us, for the first time, to address multimeric effects at correlated ab initio levels using large basis sets. At the LT-SOS-RVS-ADC(2)/def2-TZVP level, we obtain vertical excitation energies (VEEs) of 2.00-2.07 and 1.52-1.62 eV for the P680/P700 and the P870 pigment models, respectively, which agree well with the experimental absorption maxima of 1.82, 1.77, and 1.43 eV for P680, P700, and P870, respectively. In the P680/P870 models, we find that the photoexcitation leads to a π → π* transition in which the exciton is delocalized between the adjacent Chl/BChl molecules of the central pair, whereas the exciton is localized to a single chlorophyll molecule in the P700 model. Consistent with experiments, the calculated excitonic splittings between the central pairs of P680, P700, and P870 models are 80, 200, and 400 cm(-1), respectively. The calculations show that the electron affinity of the radical cation of the P680 model is 0.4 V larger than for the P870 model and 0.2 V larger than for P700. The chromophore stacking interaction is found to strongly influence the electron localization properties of the light-absorbing pigments, which may help to elucidate mechanistic details of the charge separation process in type I and type II photosystems.
View details for DOI 10.1021/acs.jctc.6b00237
View details for Web of Science ID 000378016000013
View details for PubMedID 27153186
Coupled-Cluster Studies of Extensive Green Fluorescent Protein Models Using the Reduced Virtual Space Approach
JOURNAL OF PHYSICAL CHEMISTRY B
2015; 119 (7): 2933–45
Accurate predictions of photoexcitation properties are a major challenge for modern methods of theoretical chemistry. We show here how approximate coupled-cluster singles and doubles (CC2) calculations in combination with the reduced virtual space (RVS) approach can be employed in studies of excited states of large biomolecular systems. The RVS-CC2 approach is used for accurately predicting optical properties of the p-hydroxybenzylidene-dihydroimidazolinone (p-HBDI) chromophore embedded in green fluorescent protein (GFP) models using quantum mechanical calculations in combination with large basis sets. We study the lowest excited states for the isolated and protein-embedded chromophore in two different protonation states, and show how omitting high-lying virtual orbitals in the RVS calculation of excitation energies renders large-scale CC2 studies computationally feasible. We also discuss how the error introduced by the RVS approach can be systematically estimated and controlled. The obtained CC2 excitation energies of 3.13-3.27 and 2.69-2.77 eV for the two protonation states of different protein models are in excellent agreement with the maxima of the experimental absorption spectra of 3.12-3.14 and 2.61-2.64 eV, respectively. Thus, the calculated energy splitting between the excited states of the two protonation states is 0.44-0.52 eV, which agrees very well with the experimental value of 0.48-0.51 eV. The calculations at the RVS-CC2 level on the protein models show the importance of using large QM regions in studies of biochromophores embedded in proteins.
View details for DOI 10.1021/jp5120898
View details for Web of Science ID 000349942300017
View details for PubMedID 25613980
The role of solvent exclusion in the interaction between D124 and the metal site in SOD1: implications for ALS
JOURNAL OF BIOLOGICAL INORGANIC CHEMISTRY
2013; 18 (8): 931–38
Structural changes in the metal site of the copper-zinc superoxide dismutase (SOD1) are involved in the various mechanisms proposed for the pathogenesis of the SOD1-linked familial form of amyotrophic lateral sclerosis (ALS). Elucidating how the metal site of SOD1 can be disrupted by ALS-linked mutations is important for a better understanding of the pathogenesis of the disease and for developing more efficient treatments. Residue D124, a second-sphere ligand of the copper and zinc ions, is known from experimental studies to be essential for the integrity of the metal-site structure. In this work, we used density functional theory calculations and molecular dynamics simulations to elucidate which factors keep D124 attached to the metal site and how structural changes may disrupt the binding between D124 and the metal first-sphere ligands. The calculations show that D124 is kept attached to the metal site in a kinetic trap. The exclusion of solvent molecules by the electrostatic loop of the protein is found to create the binding of D124 to the metal site. The calculations also indicate that changes in the structure of the electrostatic loop of the protein can weaken the D124-metal site interaction, lowering the affinity of the zinc site for the metal. Destabilization of the electrostatic loop of SOD1 has been previously shown to be a common property of ALS-linked variants of the protein, but its role in the pathogenesis of SOD1-linked ALS has not been elucidated.
View details for DOI 10.1007/s00775-013-1039-8
View details for Web of Science ID 000327406000006
View details for PubMedID 24026444