Department Day - Department of Chemistry

Program and abstract booklet MH Aud 7 / 30.05.2018 12:00-15:30

2018

2

PhD fellows with flash presentation and poster

Aya Ismael (p. 5): Safe chemistry to rescue carbapenems antibiotics: The last resort antibiotics

Uranbaatar Erdenebileg (p. 6): Selective photocatalytic C-H bromination of indoles

Marc Boomgaren (p. 7): Development of new anticancer inhibitors: Targeting human dUTPase

Rune Einrem (p. 8): Gold and rhenium-oxo corroles: Synthesis, spectroscopy, and applications to photodynamic therapy

Christopher Fröhlich (p. 9): Development of OXA-48 mediated ceftazidime-avibactam resistance is associated with an evolutionary trade-off

Tor Olav Berg (p. 10): From marine rest raw materials to sialic acids: Using enzymes for biomass conversion

Mikkel Christensen (p. 11): Arctic marine bacteria for bioplastic production

Typhaine Le Doujet (p. 12): Conversion of underutilized marine biomass using arctic marine bacteria as biocatalysts

Terje Klemetsen (p. 13): Evolvement of the marine reference databases

Cecilie Bækkedal (p. 14): Characterization of marine bacterial genomes and their potential for secondary metabolite production

Marc Joosten (p. 15): Four-component relativistic density functional theory calculation of electric field gradients in solids using periodic boundary conditions

Anders Brakestad (p. 16): Computational studies of an enzyme that produces bioplastics

Magnar Bjørgve (p. 17): DFT Calculations with periodic boundary conditions in a multiwavlets framework

Marc Obst (p. 18): Mechanistic study of carboxylation of alkylboranes with Cu-IPr

Ljiljana Pavlovic (p. 19): Rhodium-catalyzed hydro-carboxylation: Mechanistic analysis reveals unusual transition state for carbon–carbon bond formation

Karen Dundas (p. 20): Polarizable embedding and response theory

3

Keynote speaker

Julien S. Bourrelle (p. 4): Cracking the Norwegian Code

Poster presenter

Bjørn Olav Brandsdal (p. 21): Computation of enzyme temperature adaptation

Program

12:00-12:15 Coffee, tea, fruit & cookies

12:15-13:15 Welcome & flash presentations PhD fellows

13:15-13:30 Break

13:30-14:30 Julien S. Bourrelle: “Cracking the Norwegian Code”

14:30-15:30 Poster presentations The venue is MH building Auditorium 7 for the oral presentations and the area outside the auditorium for the poster presentations.

4

Keynote lecture: Cracking the Norwegian code Julien S. Bourrelle* Mondå Forlag, Oslo, Norway *info@monda.no "Cracking the Norwegian Code" is the lecture Julien is the most known for. It is an entertaining lecture drawing an eye-opening picture of Norwegian social behaviors and their meaning. Norwegians will learn about themselves and how their behaviors may be perceived and interpreted, while foreigners will gain valuable tools to better understand local social expectations and succeed better in a Norwegian setting. Julien's lectures use humor to make you reflect about how your culture influences you more than you think. Through entertaining stories and visual examples, you will learn about verbal and non-verbal communication, emotional feedback, social norms and values that influence interpersonal interactions across cultures. You will learn to understand better and communicate more effectively within multi-cultural environments. The lecture improves communication at work and provides practical tools to understand how to connect with people having a culture different than yours. You will learn to be more aware of your own social behaviors and how they may be misinterpreted by people of other cultures.

5

Safe Chemistry to Rescue Carbapenems Antibiotics: The Last Resort Antibiotics. Aya Ismael*1, Hanna-Kirsti S. Leiros2 and Annette Bayer1 1Department of Chemistry, UiT The Arctic University of Norway 2The Norwegian Structural Biology Centre (NorStruct), UiT The Arctic University of Norway *aya.ismael@uit.no

Figure 1: The best fragment (Kd of 9 Μm), on the left side, its crystal structure in complex with VIM-2 on the right side.

Metallo β lactamases (MBLs) are important drug target for the treatment of antibiotic resistance. Inhibitors of MBLS can restore the function of carbapenems antibiotics the last resort antibiotics for extreme antibiotic resistant infections. So far, there are no MBL inhibitors in clinical use, and hence, there is a clear need to identify novel starting points for the design of potent inhibitors. Novel fragments inhibiting the Metallo β lactamase VIM-2 have been identified [1]. The best fragment shown in Fig.1 ( Kd of 9 µM) has been chosen as the starting point for the library design [1]. The project includes the design and the synthesis of fragment library based on the fragment shown in Fig.1. In order to synthesis these type of compounds, different synthetic approaches (Scheme.1) have been evaluated. These approaches are based on Pd-carbonylative coupling reactions, which involves the handling of the toxic gas CO. To avoid the hazard of the direct use of CO, production of CO gas ex-situe using a special system (CO-ware) with a special source of CO (COgen) has been applied [2].

Scheme 1: different synthetic approaches for the targeted molecules using COgen as the CO source. References [1] Christopeit, T., Carlsen, T. J. O., Helland, R., & Leiros, H.-K. S. (2015). Journal of Medicinal Chemistry, 58(21), 8671–8682. http://doi.org/10.1021/acs.jmedchem.5b01289 [2] Hermange, P., Lindhardt, A. T., Taaning, R. H., Bjerglund, K., Lupp, D., & Skrydstrup, T. (2011). http://doi.org/10.1021/ja200818w

IBr

OO OR

Br

Br

OR

COOH

ROOCOR

HOOCO

R

CO 1.5eq,PdCl2, K2CO3

anisole110 oC, 24h

anisole110 oC, 24h

Xantphos 1-5%n-BuOH 1-2ml Pd(dba)2 5%

Xantphos 5%MePh2SiCO2H 1.5eqTMS-OK 2eq

Dioxane, 40, 2h

CO 2.5eq,PdCl2, K2CO3

CO 2.5eq , DABO boronatepd(acac)2 5%CatacXiumA.HI 10%

Toluene:H2O10:1, 80 oC, 16h

CO 1.5eqpd(dba)2 5%, DiPrPF 6%

THF, 70 oC16-20h

6

Selective photocatalytic C-H bromination of indoles Uranbaatar Erdenebileg* and Jørn H. Hansen

Department of Chemistry, UiT The Arctic University of Norway *uranbaatar.erdenebileg@uit.no Visible-light photocatalysis has recently been emerged as a powerful tool for C-H functionalization of heterocycles.1 Recently, we have published our research results regarding double C-H functionalization of indoles.2 Inspired by the formation of side-products during this study, we have developed a method of selective C-H bromination of indole derivatives using hydrobromic acid as the bromine source. We became interested in developing this transformation and herein further explore the catalytic reaction by changing reaction conditions and investigating the scope.

References [1] Douglas, J. J.; Sevrin, M. J.; Stephenson, C. R. J. Org. Process Res. Dev. 2016, 20, 1134 [2] Erdenebileg, U.; Demissie, T. B.; Hansen, J. H. Synlett 2017, 28, 907

N

R

X

N

R

XIr

HBrBr

Visible

light

(61 - 97%)

7

Development of new anticancer inhibitors: Targeting human dUTPase Marc Boomgaren*1, Marcin Miroslaw Pierechod2, Birta Ravdna Sarre1, Tor Arne Andberg3, Bjørn Olav Brandsdal3, Johan Isaksson1 and Jørn H. Hansen1 1Department of Chemistry, UiT The Arctic University of Norway 2The Norwegian Structural Biology Centre (NorStruct), Department of Chemistry, UiT The Arctic University of Norway 3Hylleraas Centre for Quantum Molecular Sciences, Department of Chemistry, UiT The Arctic University of Norway *marc.boomgaren@uit.no The inhibition of thymidylate synthase (TS) has an important role in anticancer treatment. Approved drug classes like fluoropyrimidines and antifolates leads in a depletion of the essential DNA building block deoxythymidine triphosphate (dTTP) and results among others in thymidineless death of tumor cells. [1] Unfortunately, TS-inhibitors are faced with a number of tumor resistances reducing their efficiency. [2] A key enzyme for fighting resistance is the DNA repair enzyme human deoxyuridine triphosphatase (dUTPase). It bypasses the induced cytotoxic accumulation of deoxyuridine- and 5-fluoro-2′-deoxyuridine 5′-triphosphate (dUTP and FdUTP) resulting in decreased misincorporation and apoptosis of tumor cells. [2] The inhibition of human dUTPase has the potential to “back”-boost the efficiency of approved anticancer drugs and thus makes this enzyme an interesting drug target. [1-2] We will present our latest design and synthesis strategies for new human dUTPase inhibitors based on the general template shown below and disclose our first in vitro screening results of our compounds.

References [1] Wilson P.; Danenberg P.; Johnston P.; Lenz H.; Ladner R. Nature Reviews Clinical Oncology 2014, 11, 282. [2] Miyahara, S. et al.; M. J. Med. Chem. 2012, 55, 2970.

8

Gold and rhenium-oxo corroles: Synthesis, spectroscopy, and applications to photodynamic therapy. Rune F. Einrem*1, Abraham B. Alemayehu,1 Odrun A. Gederaas2 and Abhik Ghosh1 1Department of Chemistry, UiT The Arctic University of Norway 2Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology *rune.einrem@uit.no A recent development in corrole chemistry has been the synthesis of 5d metallocorroles, which are unusual in that they combine a large metal ion with a sterically constricted macrocyclic ligand. These complexes have been found to exhibit near-IR phosphorescence ar room temperature, with low quantum yields ~1% but relatively high triplet lifetimes (~100 ms). Of these complexes, amphiphilic Au corroes showed early promise as photosensitizers in both dye-sensitized solar cells, and photodynamic therapy (PDT) experiments, which led us to explore the use of other 5d metallocorroles, particularly ReVO corroles, in similar experiments. Amphiphilic ReVO corroles exhibited high photo toxicity toward AY27 rat bladder cancer cells and WiDr human colon cancer cells. We believe that the high activity of 5d metallocorroles in these experiments reflects effective singlet oxygen production as a result of the long triplet lifetimes of these molecules.

References [1] Alemayehu, A. B. Day, N. U. Mani, T. Rubine, A. B. Thomas, K. E. Gederaas, O. A. Vinogradov, S. A. Wamser, C. C. Ghosh, A. ACS Appl. Mat. 2016 [2] Einrem, R. F. Gagnon, K. J. Alemayehu, A. B. Ghosh, A. Chem. Eur. J, 2016 [3] Einrem, R. F. Braband, H. Rof, T. Vazquez-Lima, H. Alberto, R. Ghosh, A. Chem. Eur. J., 2016

9

Development of OXA-48 mediated ceftazidime-avibactam resistance is associated with an evolutionary trade-off

Christopher Fröhlich*1, Vidar Sørum2, Hanna-Kirsti S. Leiros1, Pål Johnsen2 and Ørjan Samuelsen2,3

1Department of Chemistry, UiT The Arctic University of Norway 2Department of Pharmacy, UiT The Arctic University of Norway 3Department of Microbiology and Infection Control, University Hospital of North Norway, Tromsø, Norway *christopher.frohlich@uit.no Background: The combination of ceftazidime-avibactam (CAZ-AVI) has proven to be effective against carbapenemase-producing Gram-negative pathogens. AVI inhibits class A and some class D serine carbapenemases including OXA-48. In contrast to other carbapenemases, OXA-48 does not hydrolyse CAZ efficiently. However, OXA-48-producing isolates are frequently CAZ and multidrug-resistant due to the presence of ESBLs. The combination of CAZ-AVI is therefore likely to be used clinically on OXA-48-producing isolates. In this study, we investigated the in vitro selection of CAZ and CAZ-AVI resistance in E. coli with OXA-48. Materials/methods: In vitro resistance to CAZ and CAZ-AVI (ratio 4:1) was developed in E. coli MG1655 harboring a clinical blaOXA-48-carrying plasmid up to 64x the ancestral MIC. Point mutations within blaOXA-48 were analysed by Sanger sequencing. BlaOXA-48 and the mutated blaOXA-48 genes were cloned into pCR-Blunt II-TOPO vector and transformed into E. coli TOP10. Broth microdilution was used for MIC determination. Results: Clinical resistance to both CAZ and CAZ-AVI developed rapidly in E. coli MG1655 harboring the blaOXA-48 plasmid. DNA sequencing of blaOXA-48 revealed a single (P68A) and double mutation (P68A, Y211S) in blaOXA-48 after selection on CAZ and CAZ-AVI, respectively. Cloning of the mutated genes showed that the P68A mutation caused an 8-fold increase in CAZ MIC, but no change in the CAZ-AVI MIC. In contrast, the P68A, Y211S caused an 16-fold and 4-fold increase in CAZ and CAZ-AVI MIC, respectively. Interestingly, the mutations in OXA-48 resulted in reduced activity towards other ß-lactams (Figure 1). The MICs of piperacillin-tazobactam, temocillin, meropenem, imipenem and ertapenem all decreased by 4 to 32-fold compared to the cloned native OXA-48. Conclusions: In conclusion, our in vitro data suggest that the introduction of CAZ-AVI into clinical use could lead to OXA-48-variants with increased activity towards CAZ and reduced inhibition by AVI. However, reduced susceptibility towards CAZ-AVI comes with a trade-off, as the OXA-48 mutants show lower activity towards other ß-lactams including carbapenems. This re-sensitisation could in principle provide a therapeutic alternative.

Figure 1: Heat map of log fold-changes to different antibiotics compared to the non-mutated pCR-Blunt II-OXA-48

10

From marine rest raw materials to sialic acids: Using enzymes for biomass conversion Tor Olav Berg*, Marie-Josée Haglund Halsør, Man Kumari Gurung, Bjørn Altermark, Ronny Helland, Ulli Rothweiler, Ingar Leiros and Inger Lin U. Ræder The Norwegian Structural Biology Centre (NorStruct), Department of Chemistry, UiT The Arctic University of Norway *tor.o.berg@uit.no Sialic acids (Sias) are 9-carbon backbone sugars found on the surface of both prokaryotic and eukaryotic cells. The most well known Sia, neuraminic acid, contains an N-acetyl modification at the 5-carbon position (Neu5Ac). The Sias are involved in the recognition and mediation of cellular adhesion processes, and different variants have the potential as anti-microbial and anti-viral drugs. Neu5Ac is currently in demand, and both extraction from natural sources and industrial production give low yields or use expensive precursors. N-acetylneuraminic acid lyase (NAL) is able to synthesize Neu5Ac in a reversible reaction with the use of N-acetylmannosamine (ManNAc) and pyruvate. The enzyme is part of the sialic acid metabolism and is responsible for catabolizing Sias as a source for carbon. Several functional groups are found in nature that the enzyme has to be able to accommodate, depending on the modified carbon this will affect the enzymes affinity and ability to effectively cleave the molecule. The project involves two NALs from a marine Vibrio sp. originating from our in-house strain collection. One of the enzymes has a high percentage sequence identity to a previously characterized NAL from A. salmonicida, the other has lower sequence identity to both the others, and is suspected to use modified substrates. By analyzing the function and structure of these two enzymes it will be possible to engineer NALs with different substrate affinities in order to supply researcher and industry with a variety of Sias.

11

Arctic marine bacteria for bioplastic production Mikkel Christensen*1, Piotr Jablonski2, Seila Pandur1, Netsanet G. Assefa1, Bjørn Altermark1 and Hilde Hansen1 1The Norwegian Structural Biology Centre (NorStruct), Department of Chemistry, UiT The Arctic University of Norway 2Department of Chemistry, Umeå University, Umeå, Sweden *mikkel.christensen@uit.no Oil-based plastic production now exceeds 330 Mt annually1 and poses a major environmental problem. Polyhydroxyalkanoates (PHA) is a group of bioplastic polyesters that can replace oil-based plastics in many applications, but economic costs are higher and industrial-scale production still limited2. PHA is produced naturally by diverse groups of bacteria and stored intracellularly as hydrophobic granules, carbonosomes3, when carbon is in excess relative to other nutrients. PHA polymerase enzymes located peripherically on the carbonosome with regulatory proteins and PHA de-polymerases, synthesize the carbonosome which can occupy almost all of the cell volume for some species. We screen a collection of arctic marine bacteria to find novel PHA producers with the potential of utilizing low-value marine biomass as feedstock in production. PHA producers are selected based on fluorescence microscopy, Fourier-Transform Infra-Red (FTIR) spectral analysis and Gas Chromatography–Mass Spectrometry (GC/MS). Structural conformation of PHA monomers will be characterized using Nuclear Magnetic Resonance-spectroscopy (NMR). Growth conditions for selected bacteria is optimized to increase natural PHA production. Metabolic modelling using transcriptomic data will provide targets for metabolic engineering, eg. by knock-out or knock-down of PHA de-polymerases, to further increase PHA production. We present here results from 16s rDNA analysis of bacteria isolated from a Norwegian fish-landing facility and preliminary results of possible PHA producers from our arctic marine bacterial collection.

Figure 1: Arctic marine bacteria grown on agar plates with complex artificial seawater and additional carbon source plus the lipophilic dye NileRed, making potential PHA producing strains fluorescent under UV light (white arrows). References

[1] Statista 2018, https://www.statista.com/statistics/282732/global-production-of-plastics-since-1950/ 29.04.2018 [2] Anjum A et al. (2016). Int J Biol Macromol 89:161-174. [3] Bresan S et al. (2016). Scientific Reports 6:26612.

12

Conversion of underutilized marine biomass using arctic marine bacteria as biocatalysts Typhaine Le Doujet*, Terje Klemetsen, Concetta De Santi, Erik Hjerde, Nils P. Willassen and Peik Haugen The Norwegian Structural Biology Centre (NorStruct), Department of Chemistry, UiT The Arctic University of Norway *typhaine.l.doujet@uit.no Norway is the world’s second largest exporter of seafood with a total production of 3.28 tons in 2016. Processing of the raw material typically results in production of a main product (e.g., fish fillets, shrimp muscles), and lower value co-streams (0.91 tons in 2016). In this project, the goal is to convert underutilized co-streams to higher value products, by using microbes or enzymes as biocatalysts. In one approach, we use co-streams as feedstock in fermentation of marine microorganisms. Species are selected from a collection of bacteria from the gut microbial flora of Northeast Atlantic cod. The gut microflora represents an interesting resource because the Atlantic Cod is a generalist, which suggest that its gut microbiome must contribute to effectively break down different kinds of fish, and other marine organisms (>150 species). This is highly relevant for this project because we are using marine biomass as feedstock. Bacteria that can grow effectively on co-streams and persist as stable biofilms on plastic biobeads are of a particular interest. Finally, both single and multi-species biofilm cultures will be investigated for their potential to produce interesting molecules using both a metagenome and metabolome approach. If successful, the outcome of the project will be multiple biocatalysts that can transform cheap marine biomass to higher value products.

13

Evolvement of the Marine Reference Databases Terje Klemetsen*, Inge A. Raknes, Juan Fu, Alexander Agafonov, Sudhagar V. Balasundaram, Giacomo Tartari, Espen Robertsen and Nils P. Willassen Center for Bioinformatics (SfB), Department of Chemistry, UiT The Arctic University of Norway *terje.klemetsen@uit.no

The Marine Reference Databases (The MAR-databases) represents a collection of sequenced genomes and metagenomes originating from the marine domain. The combined lack of contextual data for samples and filtering functionality in primary databases act as a bottleneck for researchers interested in marine prokaryotes. Hosted at the Marine Metagenomics Portal (MMP) (https://mmp.sfb.uit.no/databases/), the MAR-databases; MarRef, MarDB and MarCat, which respectively hosts reference genomes of highest quality, generally sequenced genomes and a gene catalogue of proteins derived from metagenomes, all from the marine domain.

We strive to ensure our data is of marine origin and sufficiently enriched in contextual information by the means of manual curation, post-processing and post-analysis. This implies collecting material from publications and available databases, crosschecking dubious information, refining data consistency, applying defined standards and performing genome annotation and analysis of secondary metabolites using antiSMASH. The resulting metadata are freely and intuitively accessed via MMP yet supported by powerful search and filtering functionality. MMP naturally supports BLAST services and the commonly known features for querying against its sequence data.

The MAR-databases has the goal of being updated with the latest samples bi-annually, following new standards and improvements in the field as they become recognized. The latest development included the concept of Metagenome Assembled Genomes (MAG) and was implemented in the 2018 spring update (2. version) of the databases, doubling the number of entries in MarDB. The increased size required action to improve performance keeping the user experience at its best, thus the website backend has been updated with respect to data storage and compression, enabling undisturbed focus on marine sequenced data for promoting research and innovation in the field.

References

[1] Terje Klemetsen, Inge A Raknes, Juan Fu, Alexander Agafonov, Sudhagar V Balasundaram, Giacomo Tartari, Espen Robertsen, Nils P Willassen (2017): "The MAR databases: development and implementation of databases specific for marine metagenomics", Nucleic Acids Research, Volume 46, Issue D1, 4 January 2018, Pages D692–D699, https://doi.org/10.1093/nar/gkx1036

14

Characterization of marine bacterial genomes and their potential for secondary metabolite production Cecilie Bækkedal*1, Terje Klemetsen1, Marcin Miroslaw Pierechod1, Jenny Söderberg1, Concetta De Santi1, Tim Kahlke2, Erik Hjerde1, Kåre Olav Holm1, Nils Peder Willassen1, Peik Haugen1

1Center for bioinformatics (SfB) and The Norwegian Structural Biology Centre (NorStruct), Department of Chemistry, UiT The Arctic University of Norway 2Climate Change Cluster, University of Technology Sydney *cecilie.bakkedal@uit.no Marine microbes have a huge potential to produce bioactive compounds that can be beneficial for humans. In the North Atlantic Ocean, the second most abundant group of bacteria is gamma-proteobacteria. In order to investigate the potential of gamma-proteobacteria as producers of secondary metabolites, a broader understanding of the bacteria at a genomic level is needed. Pan genome analysis are performed on complete genomes from four of the most abundant orders of gamma-proteobacteria; Alteromonadales, Oceanospirillales, Chromatiales and Vibrionales. This analysis reveals species- and family-specific genes as well as unique genes. Transcriptomics are used to investigate gene expression of the species- and family-specific genes. Seven complete Vibrio anguillarum genomes have recently been published along with basic genome comparisons of the in total eleven complete V. anguillarum genomes that is publicly available [1]. This lays the groundwork for bioinformatic analysis on secondary metabolite clusters in V. anguillarum. In addition, metabolic and taxonomic profiling of multispecies biofilm communities of marine bacteria is performed. The resulting data from this project will give a better understanding of the genomics and dynamics of marine bacteria.

Figure 1: Dominating bacteria in the North Atlantic Ocean. References [1] Kåre Olav Holm, Cecilie Bækkedal, Jenny Johansson Söderberg and Peik Haugen (2018): “Complete genome sequences of seven Vibrio anguillarum strains as derived from PacBio sequencing”, Genome Biology and Evolution, evy074, https://doi.org/10.1093/gbe/evy074

15

Four-component relativistic density functional theory calculation of electric field gradients in solids using periodic boundary conditions Marc Joosten*, Marius Kadek, Michal Repisky and Kenneth Ruud

Hylleraas Centre for Quantum Molecular Sciences, Department of Chemistry, UiT The Arctic University of Norway *marc.joosten@uit.no Nuclear quadrupole interactions play an important role in various spectroscopic processes including Nuclear Magnetic Resonance and Mössbauer spectroscopies and the precise determination of the nuclear quadrupole moments and Electric Field Gradients (EFG) at the nuclei is needed for the correct interpretation of the resulting spectra. However, the theoretical prediction of inner-core properties, like the EFG, is a challenging task, because these properties are strongly influenced by relativistic effects. The following work is concerned with the calculation of Electric Field Gradients in solids using Four-Component Relativistic Density Functional Theory within periodic boundary conditions and its implementation in the ReSpect computer program. The underlying spectroscopic theory is discussed and first results are presented.

16

Computational studies of an enzyme that produces bioplastics Anders Brakestad*, Bjørn Olav Brandsdal and Kathrin H. Hopmann Hylleraas Centre for Quantum Molecular Sciences, Department of Chemistry, UiT The Arctic University of Norway *anders.m.brakestad@uit.no A large number of bacteria naturally produce long chains of polyesters called polyhydroxyalkanoates (PHA), which they use for energy storage. In addition to being biodegradable and biocompatible, PHAs also have similar properties to the widely used petroplastics, making PHAs promising candidates to replace them. However, producing PHAs is too expensive to compete with petroplastics, mainly due to the cost of raw material. The enzyme responsible for the polymerization is called “PHA synthase” (EC 2.3.1-), and in recent years three crystal structures have been published [1]–[3]. The active site of PHA synthases contains a catalytic triad comprising Asp480, His508, and Cys319 (see Fig. 1), and binds the substrate to Cys319 using covalent catalysis. With the crystal structure available, we aim to study this enzyme’s (proposed) reaction mechanism and its catalytic effect using various computational chemistry methods, such as density functional theory, molecular docking, molecular dynamics, and empirical valence bond theory. By doing this, we aim to gain a fundamental understanding of how this enzyme functions, and to suggest possible improvements to the enzyme structure to make it more effective, which may contribute to lowering the production cost of PHAs.

Figure 1: The active site of PHA synthase from PDB-code 5HZ2 [1]. Asp480, His508, and Cys319 together form the catalytic triad. His508 will abstract the H+ from Cys319, which activates Cys319 for a nucleophilic attack on the substrate. Asp480 stabilizes the positive charge on the histidine ring.

References [1] J. Kim, Y.-J. Y. Kim, S. Y. Choi, S. Y. Lee, and K.-J. K. Kim, Biotechnology Journal,

vol. 12, no. 1, p. 1600648–n/a, 2017. [2] E. C. Wittenborn, M. Jost, Y. Wei, J. A. Stubbe, and C. L. Drennan, Journal of Biological

Chemistry, vol. 291, no. 48, pp. 25264–25277, 2016. [3] M. F. Chek et al., Scientific Reports, vol. 7, no. 1, pp. 1–15, 2017.

17

DFT Calculations with periodic boundary conditions in a multiwavlets framework Magnar Bjørgve*, Stig Rune Jensen and Luca Frediani Hylleraas Centre for Quantum Molecular Sciences, Department of Chemistry, UiT The Arctic University of Norway *magnar.bjorgve@uit.no The MRChem group has for the last decade developed a code for quantum chemical calculations baseed on the MultiWavelets (MWs) basis set. This has proven to give unprecedented accuracy in molecular calculations. The objective of this project is to expand the MRChem code to solve the Kohn-Sham equations for periodic systems in order to achieve high accuracy calculations in the modelling of materials and their properties.

18

Mechanistic study of carboxylation of alkylboranes with Cu-IPr Marc F. Obst* and Kathrin H. Hopmann Hylleraas Centre for Quantum Molecular Sciences, Department of Chemistry, UiT The Arctic University of Norway *marc.obst@uit.no The mechanism of Cu-IPr-catalysed carboxylation of an alkylborane (Fig. 1) was investigated using density functional theory (PBE-D3BJ/IEFPCM). The reaction is part of a hydroboration-carboxylation protocol published by Skrydstrup et al. [1] converting a number of substituted olefins and primary alkynes into the corresponding carboxylic acids. The computed results support the proposed catalytic cycle (Fig. 1) and show two interesting aspects of the catalytic cycle. First, the transfer of the alkane from the borane to the copper occurs through a three-membered transition state (TS) involving copper, carbon and boron. Second, the preferred TS for the formation of the C-CO2 bond shows no interaction between CO2 and copper (Fig.1). This observation is in contrast to literature reports, which generally propose Cu-CO2 interactions at the TS, except for Cu-catalyzed carboxylation of sp-hybridized substrates [2]. Our recent work on Rh-catalysed hydrocarboxylations also indicates that CO2-metal interactions may not be essential [3].

Figure 1: Scheme of the carboxylation of the alkylborane (left) and TS for CO2-insertion (right), based on [1] and [4]. Distances in Å. References [1] M. Juhl, S. Laursen, ACF Catal. 2017, 7, 1392-1396. [2] M.Obst, L. Pavlovic, J. Organomet. Chem. 2018, 115-127 [3] L. Pavlovic, J. Vaitla, Organometallics 2018, 37, 941-948 [4] M. Obst, K. Hopmann, Unpublished Results.

19

Rhodium-Catalyzed Hydrocarboxylation: Mechanistic Analysis Reveals Unusual Transition State for Carbon–Carbon Bond Formation

Ljiljana Pavlovic*1, Janakiram Vaitla2, Annette Bayer2 and Kathrin H. Hopmann1 1Hylleraas Centre for Quantum Molecular Sciences, Department of Chemistry, UiT The Arctic University of Norway 2Department of Chemistry, UiT The Arctic University of Norway

*ljiljana.pavlovic@uit.no The mechanism of rhodium-COD-catalyzed hydrocarboxylation of styrene-derivatives and α,β-unsaturated carbonyl compounds with CO2 has been investigated using density functional theory (PBE-D2/IEFPCM).1 The calculations support a catalytic cycle as originally proposed by Mikami and coworkers including β-hydride elimination, insertion of the unsaturated substrate into a rhodium-hydride bond and subsequent carboxylation with CO2.2 The CO2 insertion step is found to be rate-limiting. The calculations reveal two interesting aspects: Firstly, during C-CO2 bond formation, the CO2 molecule interacts with neither the rhodium complex nor the organozinc additive. This appears to be in contrast to other CO2 insertion reactions, where CO2-metal interactions have been predicted. Secondly, the substrates show an unusual coordination mode during CO2 insertion, with the nucleophilic carbon positioned up to 3.6 Å away from rhodium. In order to understand the experimentally observed substrate preferences, we have analyzed a set of five alkenes: an α,β-unsaturated ester, an α,β-unsaturated amide, styrene and two styrene-derivatives. The analysis of the free energies shows that the ester has the lowest barrier (14.4 kcal/mol), whereas the highest activation energy was found for the amide (19.8 kcal/mol). This indicates that the barrier for CO2 insertion could explain why esters are the preferred substrates and amides are unreactive. The overall insights may be relevant for the design of future hydrocarboxylation catalysts.

References [1] Pavlovic, Ljiljana; Vaitla, Janakiram; Bayer, Annette; Hopmann, Kathrin Helen. “Rhodium-Catalyzed Hydrocarboxylation: Mechanistic Analysis Reveals Unusual Transition State for Carbon–Carbon Bond Formation.” Organometallics 2018. ISSN 0276-7333. DOI: 10.1021/acs.organomet.7b00899 [2] Kawashima, S.; Aikawa, K.; Mikami, K. “Rhodium-Catalyzed Hydrocarboxylation of Olefins with Carbon Dioxide.” Eur. J. Org. Chem. 2016, 3166-3170.

20

Polarizable embedding and response theory Karen Dundas*, Kenneth Ruud, Magnus Ringholm, Maarten Beerepoot and Jógvan Magnus Olsen Hylleraas Centre for Quantum Molecular Sciences, Department of Chemistry, UiT The Arctic University of Norway *karen.o.dundas@uit.no Within theoretical chemistry, the combination of quantum mechanical (QM) and classical methods have been used in many different forms in order to reduce computational demands while at the same time retaining accuracy. The idea is that in certain systems, some parts will be of much greater importance to the property we are studying than others, and it is therefore more important to focus on this part. This can be a solvated molecule or a functional group in a biomolecule and so on. One of these methods is Polarizable Embedding where a central part is treated with QM methods while the surrounding environment is modelled classically by placing a set of permanent multipoles and polarizabilities on the position of each atom. In this way you can model classically both electrostatic and induced interactions with the QM region. One of the many applications of theoretical chemistry is spectroscopy, and more specifically vibrational spectroscopy. Molecular properties like this are found through response theory, a name that makes sense given that the property is a response to some external influence. The neat aspect of response theory is that it shows how molecular properties can be written as derivatives of the molecular energy. For vibrational motion, the derivative is with respect to nuclear position. The fact that vibrational motion and other molecular properties are observed also in large systems, makes the combination of these two theories highly useful. Separate programs exist for each, and the job now is to combine them. When this is done, the user will be able to run efficient calculations on solutions, biomolecules or other large systems and obtain vibrational properties related to spectroscopies such as IR and Raman.

21

Computation of Enzyme Temperature Adaptation Geir Villy Isaksen1, Johan Åqvist2 and Bjørn Olav Brandsdal*1 1Hylleraas Centre for Quantum Molecular Sciences, Department of Chemistry, UiT The Arctic University of Norway 2Department of Cell and Molecular Biology, Uppsala University, Biomedical Center, Box 596, SE-751 24 Uppsala, Sweden *bjorn-olav.brandsdal@uit.no Survival of organisms in extreme environments has required expression of proteins adapted to function under a wide variety of temperatures, pH, pressure and ionic strengths. Survival in cold environments, close to the freezing point of water, renders it necessary to express heat-labile enzymes possessing high specific activity and catalytic efficiency at low temperatures. A remarkable universal characteristic of cold-active enzymes is that they show a reduction in both the activation enthalpy and entropy, compared to their warm-active orthologs, which makes their reaction rates less sensitive to decreasing temperature. The structural origin of this effect has remained largely unanswered, despite significant efforts since the early 1970s. We have investigated the temperature dependence of the reactions rates with extensive molecular dynamics free energy simulations for several warm- and cold-adapted enzyme orthologs (trypsin, endonuclease, TIM, elastase). The calculations quantitatively reproduce the catalytic rates of the enzymes and further yield high-precision Arrhenius plots, which show the characteristic trends in activation enthalpy and entropy. We find that the origin of this effect is not localized to the active site, but rather to the outer regions of the protein, where the cold-active enzyme has a higher degree of softness. Furthermore, with trypsin we show that gradually freezing the outermost parts of the enzyme causes the remarkable effect of tuning the cold-active enzyme into a variant with mesophilic characteristics without changing the amino acid sequence. Importantly, changes in the activation enthalpy – entropy balance of up to 10 kcal/mol are almost perfectly balanced at room temperature.

Figure: Warm- and cold-active enzymes have rigid cores and soft surface regions, but the cold adapted enzyme surface is softer.