Kurt Kristiansen - Academia.edu (original) (raw)

Papers by Kurt Kristiansen

5-Hydroxytryptamine 2 (serotonin 2 , 5-HT 2)-family receptors are important for mediating many ph... more 5-Hydroxytryptamine 2 (serotonin 2 , 5-HT 2)-family receptors are important for mediating many physiological functions, including vascular and nonvascular smooth muscle contraction, platelet aggregation, modulation of perception, mood, anxiety, and feeding behavior. A large number of psychopharmaceuticals, including atypical antipsychotic drugs, antidepressants, anxiolytics, and hallucinogens, mediate their actions, at least in part, via interactions with various 5-HT 2-family receptors. This review article summarizes information about structure-function aspects of 5-HT 2-family receptors. Evidence is presented that implies that conserved aromatic and charged residues are essential for ligand binding to 5-HT 2A receptors. Additionally, findings are reviewed that are consistent with the hypothesis that residues located in intracellular loops 2 and 3 (i2 and i3) mediate coupling to specific G ␣-subunits such as G ␣ q. Studies are reviewed that suggest that 5-HT 2-family receptors may be down-regulated by both agonists and antagonists, and usually this down-regulation is due to post-transcriptional mechanisms. Finally, a model for regulation of 5-HT 2-family receptors by receptor-mediated endocytosis is advanced, and the particular structural features responsible for the various endocytotic pathways are emphasized. Taken together, these results suggest that discrete domains of the receptor structure are important for ligand binding, G-protein coupling, and internalization.

Copyright information:Taken from "The third helix of the homeodomain of p... more Copyright information:Taken from "The third helix of the homeodomain of paired class homeodomain proteins acts as a recognition helix both for DNA and protein interactions"Nucleic Acids Research 2005;33(8):2661-2675.Published online 10 May 2005PMCID:PMC1092277.© The Author 2005. Published by Oxford University Press. All rights reserved The CFP- and YFP-tagged Pax6 proteins indicated in (A–E) were co-expressed in human HeLa cells. The cell images are visualized in pseudocolors before and after acceptor photobleaching to highlight changes in fluorescence intensity. Images of CFP and YFP fluorescence were obtained using the 458 nm laser line. The YFP acceptor was bleached in the whole nucleus using the 514 nm laser line. FRET was detected as decreased YFP-emission at 532 nm and a corresponding increased CFP-emission at 479 nm in the bleached area. () Full-length Pax6 proteins homodimerize in the nucleus of living cells. () The paired domain of Pax6ΔHD interacts directly with the homeodomain of the paired-less isoform Pax6ΔPD. () FRET analysis demonstrates interactions between the homeodomain of Pax6ΔPD and the paired domain of the full-length Pax6 isoform. () Mutations of arginine residues in helix 3 of the homeodomain of Pax6 impair the interaction with the paired domain, as demonstrated by no visible FRET between the HD double mutant R53A/R57A of Pax6ΔPD and the PD of full-length Pax6. () No FRET was detected between CFP and YFP in cells expressing a quadruple mutation in the paired domain of Pax6ΔHD-E101A/E112A/E120A/E128A and the wild-type homeodomain of Pax6. Cells expressing approximately a 1:1 ratio of the CFP- and YFP-tagged proteins were used for FRET experiments.

Copyright information:Taken from "The third helix of the homeodomain of p... more Copyright information:Taken from "The third helix of the homeodomain of paired class homeodomain proteins acts as a recognition helix both for DNA and protein interactions"Nucleic Acids Research 2005;33(8):2661-2675.Published online 10 May 2005PMCID:PMC1092277.© The Author 2005. Published by Oxford University Press. All rights reserved () Effect of homeodomain mutants on superactivation of Pax6 in HeLa cells. HeLa cells were co-transfected with 0.25 μg of pCI-Pax6 and 0.5 μg of either pcDNA3-HA, HA-Pax6ΔPD or HA-Pax6ΔPD mutants, together with 0.5 μg of the luciferase reporter plasmid pP6CON- and 0.05 μg of the control plasmid pCMVβ-gal. () Effect of homeodomain mutants on superactivation of Pax6ΔPD in NIH 3T3 cells. NIH 3T3 cells were co-transfected with 5 ng of Pax6ΔHD and 75 ng of either pcDNA3-HA, Pax6ΔPD or Pax6ΔPD mutants, together with 50 ng pP6CON- and 5 ng pCMV β-gal. () Reduced superactivation of the triple and the quadruple paired domain mutants of Pax6ΔHD. NIH 3T3 cells were co-transfected with 50 ng pP6CON-, 5 ng pCMVβ-gal, 5 ng of either pcDNA3-HA, Pax6ΔHD, Pax6ΔHD(E112A/E120A/E128A), Pax6ΔHD(E112A/E120A/D123A/E128A) or Pax6ΔHD(E101A/E112A/E120A/E128A) together with 75 ng pcDNA3-HA or Pax6ΔPD. () Western blot showing similar expression levels of wild type and all helix 3 mutants of Pax6ΔPD after transfection of HeLa cells. Transfection efficiencies were probed by co-transfecting an EGFP expression plasmid and developing the blot with an anti-GFP antibody. () Western blot showing similar expression levels of wild-type, triple and quadruple mutants in the PD of Pax6ΔHD. () Gel mobility shift assay with GST fusions of the PD of Pax6 wild type (WT), triple (T) and quadruple mutants (Q1 = E101A/E112A/E120A/E128A and Q2 = E112A/E120A/D123A/E128A) of Pax6 using a double-stranded oligonucleotide containing a single P6CON PD binding site as probe. The data shown (A–F) are representative of at least two other independent experiments.

Copyright information:Taken from "The third helix of the homeodomain of p... more Copyright information:Taken from "The third helix of the homeodomain of paired class homeodomain proteins acts as a recognition helix both for DNA and protein interactions"Nucleic Acids Research 2005;33(8):2661-2675.Published online 10 May 2005PMCID:PMC1092277.© The Author 2005. Published by Oxford University Press. All rights reserved () Pax6 constructs used for translation and GST pull-downs. () GST pull-down assays with Pax6 HD and PD fused to GST and immobilized on glutathione–agarose beads and Pax6ΔPDΔHD, Pax6ΔPDΔh2–3 or Pax6ΔPDΔh3 produced by transcription and translation in the presence of [S]methionine. An aliquot of 10 μl of the translation reactions was preincubated with GST immobilized on glutathione–agarose beads before incubation with the GST fusion proteins. The GST beads, GST-Pax6 HD beads and GST-Pax6 PD beads were washed several times before they were boiled in SDS loading buffer and run on a 10% SDS–polyacrylamide gel. An aliquot of 2 μl of the translated proteins was run on the same gel to visualize the signal from 20% of the input. () Point mutations in helix 3 of the homeodomain strongly reduce the ability of Pax6 HD to interact with the PD and the wild-type HD. The N51Q, R53A and R58A, but not S50A, mutants impede the HD–PD and HD–HD interactions. GST pull-down assays were performed with recombinant GST fusions of wild type or mutants of Pax6ΔHD against translated, [S]methionine-labeled Pax6, Pax6ΔHD or Pax6ΔPD. () The interactions between full-length Pax6 and the RED subdomain and between full-length Pax6 and the HD are independent of DNA. GST pull-down assays were done with Pax6 HD and RED fused to GST as in (C). Where indicated, the pull-down experiments were performed in the presence of 500 U benzonase to degrade both DNA and RNA. The results shown are representative of three independent experiments. () The PD–HD and HD–HD interactions of Pax6 are also observed in the yeast-based SOS recruitment interaction system. The temperature sensitive yeast strain MATa was co-transform [...]

be.oxfordjournals.org/ D ow nloaded from 2Abstract Examination of polymorphisms in the P. falcipa... more be.oxfordjournals.org/ D ow nloaded from 2Abstract Examination of polymorphisms in the P. falciparum gene for falcipain 2 revealed that this gene is one of two paralogs separated by 10.8 kb in chromosome 11. We designate the annotated gene denoted chr11.gen_424 as encoding falcipain 2A and that denoted chr11.gen_427 as encoding falcipain 2B. The paralogs are 96 % identical at the nucleotide level and 93 % identical at the amino acid level. The consensus sequences differ in 31/309 synonymous sites and 45/1140 nonsynonymous sites including three amino acid replacements (V393I, A400P and Q414E) that are near the catalytic site and that may affect substrate affinity or specificity. In six reference isolates, among 36 synonymous sites and 46 nonsynonymous sites that are polymorphic in the gene for falcipain 2A, falcipain 2B, or both, significant spatial clustering is observed. All but one of the polymorphisms appear to result

Computational Toxicology, 2020

Thyroid hormone disrupting chemicals (THDCs) are of major concern in ecotoxicology. With the incr... more Thyroid hormone disrupting chemicals (THDCs) are of major concern in ecotoxicology. With the increased number of emerging chemicals on the market there is a need to screen for potential THDCs in a cost-efficient way, and in silico modeling is an alternative to address this issue. In this study homology modeling and docking was used to screen a list of 626 compounds for potential thyroid hormone disrupting properties in two gull species. The tested compounds were known contaminants or emerging contaminants predicted to have the potential to reach the Arctic. Models of transthyretin (TTR) and thyroid hormone receptor α and β (TRα and TRβ) from the Arctic top predator glaucous gull (Larus hyperboreus) and temperate predator herring gull (Larus argentatus) were constructed and used to predict the binding affinity of the compounds to the thyroid hormone (TH) binding sites. The modeling predicted that 28, 4 and 330 of the contaminants would bind to TRα, TRβ and TTR respectively. These compounds were in general halogenated, aromatic and had polar functional groups, like that of THs. However, the predicted binders did not necessarily have all these properties, such as the per-and polyfluoroalkyl substances that are not aromatic and still bind to the proteins.

Computational Toxicology, 2019

Molting is an essential process in the life cycle of arthropods and is regulated by complex neuro... more Molting is an essential process in the life cycle of arthropods and is regulated by complex neuroendocrine pathways where activation of the ecdysone receptor (EcR) plays a major role. The EcR forms a non-covalent heterodimer with the ultraspiracle protein (USP) when activated by endogenous ecdysteroids, but can also be activated by several insecticides and other environmental chemicals. Environmental release of exogenous chemicals may thus represent a risk to non-target species due to phylogenetic conservation of the EcR in arthropods. In the present study, structural analysis and homology models of the EcR from the freshwater crustacean Daphnia magna were used to characterise the agonist binding pocket and identify amino acids responsible for differences in agonist binding between arthropod species. The analysis showed that the binding pockets of steroidal and non-steroidal agonists are partly overlapping, and the phylogenetically conserved Thr59 is a key residue for binding both types of agonists. In silico site-directed mutagenesis and MM-GBSA dG calculations revealed that Cys100 (D. magna numbering) is a structural determinant for cross species affinities. Other determinants are Val129 for both types of agonists, Thr132 for steroidal agonists and Asp134 for non-steroidal agonists. The present results can be used to predict cross species sensitivity for EcR agonists, and shows that homology modelling and affinity predictions may contribute to identifying susceptible species for EcR-mediated endocrine disruption.

Toxicological Sciences, 2018

Chemical hazard assessment requires extrapolation of information from model organisms to all spec... more Chemical hazard assessment requires extrapolation of information from model organisms to all species of concern. The Sequence Alignment to Predict Across Species Susceptibility (SeqAPASS) tool was developed as a rapid, cost-effective method to aid cross-species extrapolation of susceptibility to chemicals acting on specific protein targets through evaluation of protein structural similarities and differences. The greatest resolution for extrapolation of chemical susceptibility across species involves comparisons of individual amino acid residues at key positions involved in proteinchemical interactions. However, a lack of understanding of whether specific amino acid substitutions among species at key positions in proteins affect interaction with chemicals made manual interpretation of alignments time consuming and potentially inconsistent. Therefore, this study used in silico site-directed mutagenesis coupled with docking simulations of computational models for acetylcholinesterase (AChE) and ecdysone receptor (EcR) to investigate how specific amino acid substitutions impact protein-chemical interaction. This study found that computationally derived substitutions in identities of key amino acids caused no change in protein-chemical interaction if residues share the same side chain functional properties and have comparable molecular dimensions, while differences in these characteristics can change protein-chemical interaction. These findings were considered in the development of capabilities for automatically generated species-specific predictions of chemical susceptibility in SeqAPASS. These predictions for AChE and EcR were shown to agree with SeqAPASS predictions comparing the primary sequence and functional domain sequence of proteins for more than 90% of the investigated species, but also identified dramatic species-specific differences in chemical susceptibility that align with results from standard toxicity tests. These results provide a compelling line of evidence for use of SeqAPASS in deriving screening level, species-specific, susceptibility predictions across broad taxonomic groups for application to human and ecological hazard assessment.

Journal of chemical information and modeling, Jan 18, 2017

Despite its remarkable importance in the arena of drug design, serotonin 1A receptor (5-HT1A) has... more Despite its remarkable importance in the arena of drug design, serotonin 1A receptor (5-HT1A) has been elusive to the X-ray crystallography community. This lack of direct structural information not only hampers our knowledge regarding the binding modes of many popular ligands (including the endogenous neurotransmitter-serotonin), but also limits the search for more potent compounds. In this paper we shed new light on the 3D pharmacological properties of the 5-HT1A receptor by using a ligand-guided approach (ALiBERO) grounded in the Internal Coordinate Mechanics (ICM) docking platform. Starting from a homology template and set of known actives, the method introduces receptor flexibility via Normal Mode Analysis and Monte Carlo sampling, to generate a subset of pockets that display enriched discrimination of actives from inactives in retrospective docking. Here, we thoroughly investigated the repercussions of using different protein templates and the effect of compound selection on sc...

Environmental Science & Technology, 2016

We studied interactions between polar bear peroxisome proliferator-activated receptor gamma (pbPP... more We studied interactions between polar bear peroxisome proliferator-activated receptor gamma (pbPPARG) and selected compounds using a luciferase reporter assay and predictions through molecular docking. Furthermore, we studied adipogenesis by liver and adipose tissue extracts from a polar bear and three synthetic mixtures of contaminants in murine 3T3-L1 preadipocytes and polar bear adipose tissue-derived stem cells (pbASCs). PCB153 and p,p'-DDE antagonized pbPPARG, although their predicted receptor-ligand affinity was weak. PBDEs, tetrabromobisphenol A, and PCB170 had a weak agonistic effect on pbPPARG, while hexabromocyclododecane, bisphenol A, oxychlordane, and endosulfan were weak antagonists. pbPPARG-mediated luciferase activity was suppressed by synthetic contaminant mixtures reflecting levels measured in polar bear adipose tissue, as were transcript levels of PPARG and the PPARG target gene fatty acid binding protein 4 (FABP4) in pbASCs. Contaminant extracts from polar bear tissues enhanced triglyceride accumulation in murine 3T3-L1 cells and pbASCs, whereas triglyceride accumulation was not affected by the synthetic mixtures. Chemical characterization of extracts using non-target methods revealed presence of exogenous compounds that have previously been reported to induce adipogenesis. These compounds included phthalates, tonalide, and nonylphenol. In conclusion, major legacy contaminants in polar bear adipose tissue exert antagonistic effects on PPARG, but adipogenesis by a mixture containing emerging compounds may be enhanced through PPARG or other pathways.

PLoS ONE, 2013

This study explores a new approach to pharmacophore screening involving the use of an optimized l... more This study explores a new approach to pharmacophore screening involving the use of an optimized linear combination of models instead of a single hypothesis. The implementation and evaluation of the developed methodology are performed for a complete known chemical space of 5-HT 1A R ligands (3616 active compounds with K i < 100 nM) acquired from the ChEMBL database. Clusters generated from three different methods were the basis for the individual pharmacophore hypotheses, which were assembled into optimal combinations to maximize the different coefficients, namely, MCC, accuracy and recall, to measure the screening performance. Various factors that influence filtering efficiency, including clustering methods, the composition of test sets (random, the most diverse and cluster population-dependent) and hit mode (the compound must fit at least one or two models from a final combination) were investigated. This method outmatched both single hypothesis and random linear combination approaches.

Bioorganic & Medicinal Chemistry Letters, 2011

We introduce a new approach to the known concept of interaction profiles, based on Structural Int... more We introduce a new approach to the known concept of interaction profiles, based on Structural Interaction Fingerprints (SIFt), for precise and rapid binding site description. A set of scripts for batch generation and analysis of SIFt were prepared, and the implementation is computationally efficient and supports parallelization. It is based on a 9-digit binary interaction pattern that describes physical ligand-protein interactions in structures and models of ligand-protein complexes. The tool performs analysis and identifies binding site residues (crucial and auxiliary) and classifies interactions according to type (hydrophobic, aromatic, charge, polar, side chain, and backbone). It is convenient and easy to use, and gives manageable output data for both, interpretation and further processing. In the presented Letter, SIFts are applied to analyze binding sites in models of antagonist-5-HT7 receptor complexes and structures of cyclin dependent kinase 2-ligand complexes.

Receptors & channels, 1998

Molecular modeling techniques were used to build a three-dimensional model of the human 5-HT1B re... more Molecular modeling techniques were used to build a three-dimensional model of the human 5-HT1B receptor. The receptor model was used to examine receptor interactions of 5-hydroxytryptamine (serotonin), (S)pindolol and of the tetrapeptide Leu-Ser-Ala-Leu (LSAL), which recently has been shown to interact specifically with the 5-HT1B receptor. We have assumed that the NH3(+)-LSAL-COO- form of the tetrapeptide is the biologically active, and propose that a negatively charged residue conserved among various species homologues of the 5-HT1B receptor may act as a counter-ion for the positively charged N-terminus of the tetrapeptide. The strongest LSAL-receptor interactions were obtained after molecular dynamics simulations that were started with the N-terminus of LSAL positioned close to Asp352 in transmembrane helix 7. The model suggests that Asp352 in transmembrane helix 7 may act as a counter-ion for the positively charged N-terminus, and that the side chains of Tyr109 (transmembrane he...

Journal of Pharmacology and Experimental Therapeutics, Jun 1, 2000

Discovering the molecular and atomic mechanism(s) by which G-protein-coupled receptors (GPCRs) ar... more Discovering the molecular and atomic mechanism(s) by which G-protein-coupled receptors (GPCRs) are activated by agonists remains an elusive goal. Recently, studies examining two representative GPCRs (rhodopsin and ␣ 1b -adrenergic receptors) have suggested that the disruption of a putative "saltbridge" between highly conserved residues in transmembrane (TM) helix III, involving aspartate or glutamate, and helix VII, involving a basic residue, results in receptor activation. We have tested whether this is a general mechanism for GPCR activation by constructing a model of the 5-hydroxytryptamine (5-HT) 2A receptor and characterizing several mutations at the homologous residues (Asp-155 and Asn-363) of the 5-HT 2A serotonin receptor. All of the mutants (D155A, D155N, D155E, D155Q, and S363A) resulted in receptors with reduced basal

Copyright information:Taken from "The third helix of the homeodomain of p... more Copyright information:Taken from "The third helix of the homeodomain of paired class homeodomain proteins acts as a recognition helix both for DNA and protein interactions"Nucleic Acids Research 2005;33(8):2661-2675.Published online 10 May 2005PMCID:PMC1092277.© The Author 2005. Published by Oxford University Press. All rights reserved () Effect of homeodomain mutants on superactivation of Pax6 in HeLa cells. HeLa cells were co-transfected with 0.25 μg of pCI-Pax6 and 0.5 μg of either pcDNA3-HA, HA-Pax6ΔPD or HA-Pax6ΔPD mutants, together with 0.5 μg of the luciferase reporter plasmid pP6CON- and 0.05 μg of the control plasmid pCMVβ-gal. () Effect of homeodomain mutants on superactivation of Pax6ΔPD in NIH 3T3 cells. NIH 3T3 cells were co-transfected with 5 ng of Pax6ΔHD and 75 ng of either pcDNA3-HA, Pax6ΔPD or Pax6ΔPD mutants, together with 50 ng pP6CON- and 5 ng pCMV β-gal. () Reduced superactivation of the triple and the quadruple paired domain mutants of Pax6ΔHD. NIH 3T3 cells were co-transfected with 50 ng pP6CON-, 5 ng pCMVβ-gal, 5 ng of either pcDNA3-HA, Pax6ΔHD, Pax6ΔHD(E112A/E120A/E128A), Pax6ΔHD(E112A/E120A/D123A/E128A) or Pax6ΔHD(E101A/E112A/E120A/E128A) together with 75 ng pcDNA3-HA or Pax6ΔPD. () Western blot showing similar expression levels of wild type and all helix 3 mutants of Pax6ΔPD after transfection of HeLa cells. Transfection efficiencies were probed by co-transfecting an EGFP expression plasmid and developing the blot with an anti-GFP antibody. () Western blot showing similar expression levels of wild-type, triple and quadruple mutants in the PD of Pax6ΔHD. () Gel mobility shift assay with GST fusions of the PD of Pax6 wild type (WT), triple (T) and quadruple mutants (Q1 = E101A/E112A/E120A/E128A and Q2 = E112A/E120A/D123A/E128A) of Pax6 using a double-stranded oligonucleotide containing a single P6CON PD binding site as probe. The data shown (A–F) are representative of at least two other independent experiments.

Copyright information:Taken from "The third helix of the homeodomain of p... more Copyright information:Taken from "The third helix of the homeodomain of paired class homeodomain proteins acts as a recognition helix both for DNA and protein interactions"Nucleic Acids Research 2005;33(8):2661-2675.Published online 10 May 2005PMCID:PMC1092277.© The Author 2005. Published by Oxford University Press. All rights reserved () Pax6 constructs used for translation and GST pull-downs. () GST pull-down assays with Pax6 HD and PD fused to GST and immobilized on glutathione–agarose beads and Pax6ΔPDΔHD, Pax6ΔPDΔh2–3 or Pax6ΔPDΔh3 produced by transcription and translation in the presence of [S]methionine. An aliquot of 10 μl of the translation reactions was preincubated with GST immobilized on glutathione–agarose beads before incubation with the GST fusion proteins. The GST beads, GST-Pax6 HD beads and GST-Pax6 PD beads were washed several times before they were boiled in SDS loading buffer and run on a 10% SDS–polyacrylamide gel. An aliquot of 2 μl of the translated proteins was run on the same gel to visualize the signal from 20% of the input. () Point mutations in helix 3 of the homeodomain strongly reduce the ability of Pax6 HD to interact with the PD and the wild-type HD. The N51Q, R53A and R58A, but not S50A, mutants impede the HD–PD and HD–HD interactions. GST pull-down assays were performed with recombinant GST fusions of wild type or mutants of Pax6ΔHD against translated, [S]methionine-labeled Pax6, Pax6ΔHD or Pax6ΔPD. () The interactions between full-length Pax6 and the RED subdomain and between full-length Pax6 and the HD are independent of DNA. GST pull-down assays were done with Pax6 HD and RED fused to GST as in (C). Where indicated, the pull-down experiments were performed in the presence of 500 U benzonase to degrade both DNA and RNA. The results shown are representative of three independent experiments. () The PD–HD and HD–HD interactions of Pax6 are also observed in the yeast-based SOS recruitment interaction system. The temperature sensitive yeast strain MATa was co-transform [...]