Nikolina Sekulic

• Heggelund, Julie Elisabeth; Ahmad, Ahmad Ali; Johannesen, Marie Kongshaug; Sekulic, Nikolina & Sollid, Ludvig Magne (2026). Cryo-EM structure of the complex between transglutaminase 2 and the 45 kDa domain of fibronectin. Journal of Biological Chemistry. ISSN 0021-9258. 302(9), p. 1–11. doi: 10.1016/j.jbc.2026.113326. Full text in Research Archive
• Gupta, Kushol; Sekulic, Nikolina; Allu, Praveen Kumar; Sapp, Nicklas; Huang, Qingqiu & Sarachan, Kathryn [Show all 13 contributors for this article] (2025). Solution conformational differences between conventional and CENP-A nucleosomes are accentuated by reversible deformation under high pressure. Chromosome Research. ISSN 0967-3849. 33(1). doi: 10.1007/s10577-025-09769-z. Full text in Research Archive Show summary

  Abstract Solution-based interrogation of the physical nature of nucleosomes has its roots in X-ray and neutron scattering experiments, including those that provided the initial observation that DNA wraps around core histones. In this study, we performed a comprehensive small-angle scattering study to compare canonical nucleosomes with variant centromeric nucleosomes harboring the histone variant, CENP-A. We used nucleosome core particles (NCPs) assembled on an artificial positioning sequence (Widom 601) and compared these to those assembled on a natural α-satellite DNA from human centromeres. We establish the native solution properties of octameric H3 and CENP-A NCPs using analytical ultracentrifugation (AUC), small-angle X-ray scattering (SAXS), and contrast variation small-angle neutron scattering (CV-SANS). Using high-pressure SAXS (HP-SAXS), we discovered that both histone and DNA sequence have an impact on the stability of octameric nucleosomes in solution under high pressure (300 MPa), with evidence of reversible unwrapping in these experimental conditions. Both canonical nucleosomes harboring conventional histone H3 and their centromeric counterparts harboring CENP-A have a substantial increase in their radius of gyration, but this increase is much less prominent for centromeric nucleosomes. More broadly for chromosome-related research, we note that as HP-SAXS methodologies expand in their utility, we anticipate this will provide a powerful solution-based approach to study nucleosomes and higher-order chromatin complexes.

• Sankaranarayanan, Sundar Ram; Ulmer, Jonathan; Mørch, Anna; Ahmad, Ahmad Ali; Sekulic, Nikolina & Drinnenberg, Ines Anna (2025). Insects evolved a monomeric histone-fold domain in the CENP-T protein family. EMBO Reports. ISSN 1469-221X. 26(23), p. 5825–5825. doi: 10.1038/s44319-025-00603-5. Full text in Research Archive Show summary

  Abstract The histone-fold domain (HFD) is a conserved protein interaction module that requires stabilization through a handshake interaction with an HFD partner. All HFD proteins known to date form obligate dimers to shield the extensive hydrophobic residues along the HFD. Here, we find that the lepidopteran kinetochore protein CENP-T is soluble as a monomer. We attribute this stability to a structural rearrangement, which leads to the repositioning of the HFD helix α3. This brings a conserved two-helical extension closer to the histone fold, where it takes over the position and function of the CENP-T partner CENP-W. This change has no effect on the DNA-binding ability of the lepidopteran CENP-T. Our analysis suggests that the monomeric HFD originated in the last common ancestor of insects, with a possible second independent origin in Acariformes, both of which lack CENP-W. Our study highlights an unexpected structural variation in a protein module as conserved and optimized as the HFD, providing a unique perspective on the evolution of protein structure and the forces driving it.

• Timmer, Romy; Bellanger, Aurélie Nathalie Pascale; Peeters, Sarah; Kim, Hera; Ballina, Laura Rodriguez de la & Eikvar, Sissel [Show all 11 contributors for this article] (2025). LRRC59 cooperates with nuclear transporters to restrain the nuclear envelope repair machinery and safeguard genome integrity. Nature Communications. 16(1). doi: 10.1038/s41467-025-65994-4. Full text in Research Archive Show summary

  Nuclear envelope (NE) rupture is a hallmark of cancer cells, and persistent NE damage drives genome instability and inflammation. NE repair relies on activation of the endosomal sorting complex required for transport (ESCRT)-III repair machinery by the LEMD2-CHMP7 compartmentalization sensor, but little is known beyond these core factors. Here, we use convergent proximity proteomics to inventorise proteins mobilized to the NE upon assembly of LEMD2-CHMP7 and activation of ESCRT-III. Within this NE repairome, we identify LRRC59 as a critical regulator of LEMD2 accumulation at NE ruptures. We find that LRRC59, together with the nuclear transporters KPNB1 and XPO1, restricts the assembly of LEMD2-CHMP7 complexes to the site of rupture. Disruption of this regulatory axis escalates LEMD2-CHMP7 spreading across the NE, driving torsional DNA damage in ruptured nuclei and micronuclei. Thus, our work identifies a central regulatory layer of NE repair centered on LRRC59 and KPNB1. We propose that altered LRRC59 levels and deregulated nuclear transport coordinately compromise NE repair, driving genome instability and cancer development.

• Ahmad, Ahmad Ali; Mors, Mira; Carrer, Manuel; Li, Xinmeng; Bilokapić, Silvija & Halić, Mario [Show all 8 contributors for this article] (2025). Structure and dynamics of 2x(CENP-A/H4)2 octasome reveal a possible intermediate in centromeric chromatin. Life Science Alliance (LSA). 9(3). doi: 10.26508/lsa.202503377. Full text in Research Archive Show summary

  The centromere is a part of the chromosome that is essential for the even segregation of duplicated chromosomes during cell division. It is epigenetically defined by the presence of the histone H3 variant CENP-A. CENP-A associates specifically with a group of 16 proteins that form the constitutive centromere-associated network (CCAN) of proteins. In mitosis, the kinetochore forms on the CCAN to connect the duplicated chromosomes to the microtubules protruding from the cell poles. Previous studies have shown that CENP-A replaces H3 in nucleosomes, and recently, the structures of CENP-A-containing nucleosomes in complex with CCANs have been revealed, but they show only a limited interaction between CCANs and CENP-A. Here, we report the cryo-EM structure of 2x(CENP-A/H4)2 octasomes assembled on DNA in the absence of H2A/H2B histone dimer and speculate how (CENP-A/H4)2 tetrasomes might serve as a platform for CCAN organization.

• Petersson, Marcus; Zingl, Franz G.; Rodriguez-Rodriguez, Everardo; Rendsvig, Jakob K. H.; Heinsøe, Heidi & Arendrup, Emma Wenzel [Show all 16 contributors for this article] (2025). Orally delivered toxin–binding protein protects against diarrhoea in a murine cholera model. Nature Communications. 16. doi: 10.1038/s41467-025-57945-w. Full text in Research Archive Show summary

  The ongoing seventh cholera pandemic, which began in 1961, poses an escalating threat to public health. There is a need for new cholera control measures, particularly ones that can be produced at low cost, for the one billion people living in cholera-endemic regions. Orally delivered VHHs, functioning as target-binding proteins, have been proposed as a potential approach to control gastrointestinal pathogens. Here, we describe the development of an orally deliverable bivalent VHH construct that binds to the B-pentamer of cholera toxin, showing that it inhibits toxin activity in a murine challenge model. Infant mice given the bivalent VHH prior to V. cholerae infection exhibit a significant reduction in cholera toxin–associated intestinal fluid secretion and diarrhoea. In addition, the bivalent VHH reduces V. cholerae colonization levels in the small intestine by a factor of 10. This cholera toxin–binding protein holds promise for protecting against severe diarrhoea associated with cholera.

• Brugger, Melanie; Lauri, Antonella; Zhen, Yan; Gramegna, Laura L.; Zott, Benedikt & Sekulic, Nikolina [Show all 40 contributors for this article] (2024). Bi-allelic variants in SNF8 cause a disease spectrum ranging from severe developmental and epileptic encephalopathy to syndromic optic atrophy. American Journal of Human Genetics. ISSN 0002-9297. 111(3), p. 594–613. doi: 10.1016/j.ajhg.2024.02.005. Full text in Research Archive Show summary

  The endosomal sorting complex required for transport (ESCRT) machinery is essential for membrane remodeling and autophagy and it comprises three multi-subunit complexes (ESCRT I-III). We report nine individuals from six families presenting with a spectrum of neurodevelopmental/neurodegenerative features caused by bi-allelic variants in SNF8 (GenBank: NM_007241.4), encoding the ESCRT-II subunit SNF8. The phenotypic spectrum included four individuals with severe developmental and epileptic encephalopathy, massive reduction of white matter, hypo-/aplasia of the corpus callosum, neurodevelopmental arrest, and early death. A second cohort shows a milder phenotype with intellectual disability, childhood-onset optic atrophy, or ataxia. All mildly affected individuals shared the same hypomorphic variant, c.304G>A (p.Val102Ile). In patient-derived fibroblasts, bi-allelic SNF8 variants cause loss of ESCRT-II subunits. Snf8 loss of function in zebrafish results in global developmental delay and altered embryo morphology, impaired optic nerve development, and reduced forebrain size. In vivo experiments corroborated the pathogenicity of the tested SNF8 variants and their variable impact on embryo development, validating the observed clinical heterogeneity. Taken together, we conclude that loss of ESCRT-II due to bi-allelic SNF8 variants is associated with a spectrum of neurodevelopmental/neurodegenerative phenotypes mediated likely via impairment of the autophagic flux.

• Pena, Dario Segura; Hovet, Oda; Gogoi, Hemanga; Dawicki-Mckenna, Jennine; Wøien, Stine Malene Hansen & Carrer, Manuel [Show all 9 contributors for this article] (2023). The structural basis of the multi-step allosteric activation of Aurora B kinase. eLIFE. 12. doi: 10.7554/eLife.85328. Full text in Research Archive Show summary

  Aurora B, together with IN-box, the C-terminal part of INCENP, forms an enzymatic complex that ensures faithful cell division. The [Aurora B/IN-box] complex is activated by autophosphorylation in the Aurora B activation loop and in IN-box, but it is not clear how these phosphorylations activate the enzyme. We used a combination of experimental and computational studies to investigate the effects of phosphorylation on the molecular dynamics and structure of [Aurora B/IN-box]. In addition, we generated partially phosphorylated intermediates to analyze the contribution of each phosphorylation independently. We found that the dynamics of Aurora and IN-box are interconnected, and IN-box plays both positive and negative regulatory roles depending on the phosphorylation status of the enzyme complex. Phosphorylation in the activation loop of Aurora B occurs intramolecularly and prepares the enzyme complex for activation, but two phosphorylated sites are synergistically responsible for full enzyme activity.

• Nagpal, Harsh; Ahmad, Ahmad Ali; Hirano, Yasuhiro; Cai, Wei; Halic, Mario & Fukagawa, Tatsuo [Show all 8 contributors for this article] (2023). CENP-A and CENP-B collaborate to create an open centromeric chromatin state. Nature Communications. 14(1). doi: 10.1038/s41467-023-43739-5. Full text in Research Archive Show summary

  Centromeres are epigenetically defined via the presence of the histone H3 variant CENP-A. Contacting CENP-A nucleosomes, the constitutive centromere associated network (CCAN) and the kinetochore assemble, connecting the centromere to spindle microtubules during cell division. The DNA-binding centromeric protein CENP-B is involved in maintaining centromere stability and, together with CENP-A, shapes the centromeric chromatin state. The nanoscale organization of centromeric chromatin is not well understood. Here, we use single-molecule fluorescence and cryoelectron microscopy (cryoEM) to show that CENP-A incorporation establishes a dynamic and open chromatin state. The increased dynamics of CENP-A chromatin create an opening for CENP-B DNA access. In turn, bound CENP-B further opens the chromatin fiber structure and induces nucleosomal DNA unwrapping. Finally, removal of CENP-A increases CENP-B mobility in cells. Together, our studies show that the two centromere-specific proteins collaborate to reshape chromatin structure, enabling the binding of centromeric factors and establishing a centromeric chromatin state.

• Sekulic, Nikolina; Hovet, Oda; Gogoi, Hemanga; Dawicki-McKenna, Jennine; Wøien, Stine Malene Hansen & Carrer, Manuel [Show all 8 contributors for this article] (2022). The structural basis of the multi-step allosteric activation of Aurora B kinase. BioRxiv. ISSN 0362-4331. Full text in Research Archive
• Dorraji, Seyed Esmaeil; Borgen, Elin; Segura-Pena, Dario; Rawat, Puneet; Smorodina, Eva & Dunn, Claire [Show all 10 contributors for this article] (2022). Development of a High-Affinity Antibody against the Tumor-Specific and Hyperactive 611-p95HER2 Isoform. Cancers. 14:4859(19), p. 1–24. doi: 10.3390/cancers14194859. Full text in Research Archive Show summary

  Simple Summary In the present study, we addressed the unmet need for a molecular antibody (mAb) with high affinity and specificity against a truncated hyperactive isoform of human epidermal growth factor receptor 2 (HER2), called 611-carboxy terminal fragment (CTF)-p95HER2. Patients with p95HER2+ breast cancer are at risk of developing metastatic breast cancer with a poor prognosis and resistance to therapies targeting full-length HER2. We have generated a mAb named Oslo-2, which react specifically with 611-CTF-p95HER2 and has a high affinity. We also characterized the antigenic determinant (epitope) on the p95HER2 protein and the antigen-binding site (paratope) on the Oslo-2 mAb. The antibody can be used to develop antibody- or cell-based therapies targeting p95HER2, as well as a diagnostic assay to identify p95HER2+ disease. Abstract The expression of human epidermal growth factor receptor 2 (HER2) is a key classification factor in breast cancer. Many breast cancers express isoforms of HER2 with truncated carboxy-terminal fragments (CTF), collectively known as p95HER2. A common p95HER2 isoform, 611-CTF, is a biomarker for aggressive disease and confers resistance to therapy. Contrary to full-length HER2, 611-p95HER2 has negligible normal tissue expression. There is currently no approved diagnostic assay to identify this subgroup and no therapy targeting this mechanism of tumor escape. The purpose of this study was to develop a monoclonal antibody (mAb) against 611-CTF-p95HER2. Hybridomas were generated from rats immunized with cells expressing 611-CTF. A hybridoma producing a highly specific Ab was identified and cloned further as a mAb. This mAb, called Oslo-2, gave strong staining for 611-CTF and no binding to full-length HER2, as assessed in cell lines and tissues by flow cytometry, immunohistochemistry and immunofluorescence. No cross-reactivity against HER2 negative controls was detected. Surface plasmon resonance analysis demonstrated a high binding affinity (equilibrium dissociation constant 2 nM). The target epitope was identified at the N-terminal end, using experimental alanine scanning. Further, the mAb paratope was identified and characterized with hydrogen-deuterium-exchange, and a molecular model for the (Oslo-2 mAb:611-CTF-p95HER2) complex was generated by an experimental-information-driven docking approach. We conclude that the Oslo-2 mAb has a high affinity and is highly specific for 611-CTF-p95HER2. The Ab may be used to develop potent and safe therapies, overcoming p95HER2-mediated tumor escape, as well as for developing diagnostic assays. Keywords: HER2; p95HER2; antibody; tumor-specific; breast cancer; cancer stem cells; HDX-MS; epitope and paratope mapping; docking

• Stonyte, Vilte; Martîn, Ruth Martîn; Segura-Pena, Dario; Sekulic, Nikolina & Aviles, Sandra Lopez (2020). Requirement of PP2A-B56 Par1 for the Stabilization of the CDK Inhibitor Rum1 and Activation of APC/C Ste9 during Pre-Start G1 in S. pombe. iScience. 23(5). doi: 10.1016/j.isci.2020.101063. Full text in Research Archive Show summary

  Exit from the cell cycle during the establishment of quiescence and upon cell differentiation requires the sustained inactivation of CDK complexes. Fission yeast cells deprived of nitrogen halt cell cycle progression in pre-Start G1, before becoming quiescent or undergoing sexual differentiation. The CDK inhibitor Rum1 and the APC/C activator Ste9 are fundamental for this arrest, but both are down-regulated by CDK complexes. Here, we show that PP2A-B56Par1 is instrumental for Rum1 stabilization and Ste9 activation. In the absence of PP2A-B56Par1, cells fail to accumulate Rum1, and this results in persistent CDK activity, Ste9 inactivation, retention of the mitotic cyclin Cdc13, and impaired withdrawal from the cell cycle during nitrogen starvation. Importantly, mutation of a putative B56 interacting motif in Rum1 recapitulates these defects. These results underscore the relevance of CDK-counteracting phosphatases in cell differentiation, establishment of the quiescent state, and escape from it in cancer cells.

• Ahmad, Ahmad Ali & Sekulic, Nikolina (2020). CENP-A nucleosome-a chromatin-embedded pedestal for the centromere: lessons learned from structural biology. Essays in Biochemistry. ISSN 0071-1365. 64(2), p. 205–221. doi: 10.1042/EBC20190074. Full text in Research Archive Show summary

  The centromere is a chromosome locus that directs equal segregation of chromosomes during cell division. A nucleosome containing the histone H3 variant CENP-A epigenetically defines the centromere. Here, we summarize findings from recent structural biology studies, including several CryoEM structures, that contributed to elucidate specific features of the CENP-A nucleosome and molecular determinants of its interactions with CENP-C and CENP-N, the only two centromere proteins that directly bind to it. Based on those findings, we propose a role of the CENP-A nucleosome in the organization of centromeric chromatin beyond binding centromeric proteins.

• Ahmad, Ahmad Ali; Bilokapic, Silvija; Schafer, Ingmar B; Halic, Mario & Sekulic, Nikolina (2019). CENP‐C unwraps the human CENP‐A nucleosome through the H2A C‐terminal tail. EMBO Reports. ISSN 1469-221X. doi: 10.15252/embr.201948913. Full text in Research Archive Show summary

  Centromeres are defined epigenetically by nucleosomes containing the histone H3 variant CENP ‐A, upon which the constitutive centromere‐associated network of proteins (CCAN ) is built. CENP ‐C is considered to be a central organizer of the CCAN . We provide new molecular insights into the structure of human CENP ‐A nucleosomes, in isolation and in complex with the CENP ‐C central region (CENP ‐CCR ), the main CENP ‐A binding module of human CENP ‐C. We establish that the short αN helix of CENP ‐A promotes DNA flexibility at the nucleosome ends, independently of the sequence it wraps. Furthermore, we show that, in vitro , two regions of human CENP ‐C (CENP ‐CCR and CENP ‐Cmotif) both bind exclusively to the CENP ‐A nucleosome. We find CENP ‐CCR to bind with high affinity due to an extended hydrophobic area made up of CENP ‐AV 532 and CENP ‐AV 533. Importantly, we identify two key conformational changes within the CENP ‐A nucleosome upon CENP ‐C binding. First, the loose DNA wrapping of CENP ‐A nucleosomes is further exacerbated, through destabilization of the H2A C‐terminal tail. Second, CENP ‐CCR rigidifies the N‐terminal tail of H4 in the conformation favoring H4K20 monomethylation, essential for a functional centromere.

• Sekulic, Nikolina & Black, Ben E. (2016). Preparation of Recombinant Centromeric Nucleosomes and Formation of Complexes with Nonhistone Centromere Proteins. Methods in Enzymology. ISSN 0076-6879. 573, p. 67–96. doi: 10.1016/bs.mie.2016.01.014. Full text in Research Archive Show summary

  Centromeres are present on each chromosome to direct proper segregation during cell division. The understanding of how the histone H3 variant, CENP-A, epigenetically marks the location of the centromere on the chromosome has been advanced, in part, through the study of histone complexes, nucleosomes, and nucleosomal complexes with nonhistone centromere proteins. In this chapter, we describe the preparation of recombinant versions of these complexes. The methodology is firmly rooted in classic nucleosome reconstitution methods, but we highlight the aspects of the preparations that diverge from those used for the methods established with canonical histones. We also provide a method for producing PCR-amplified nucleosomal DNA sequences in milligram quantities that is particularly useful for studies where multiple sequences and/or chemical modifications are desired. Lastly, we describe our approach to assemble and analyze a complex between the recombinant human CENP-A nucleosome and one of its binding partners, CENP-C.

• Falk, Samantha J.; Lee, Jaehyoun; Sekulic, Nikolina; Sennett, Michael A.; Lee, Tae-Hee & Black, Ben E. (2016). CENP-C directs a structural transition of CENP-A nucleosomes mainly through sliding of DNA gyres. Nature Structural & Molecular Biology. ISSN 1545-9993. 23(3), p. 204–208. doi: 10.1038/nsmb.3175. Full text in Research Archive
• Avgousti, Daphne C.; Herrmann, Christin; Kulej, Katarzyna; Pancholi, Neha J.; Sekulic, Nikolina & Petrescu, Joana [Show all 17 contributors for this article] (2016). A core viral protein binds host nucleosomes to sequester immune danger signals. Nature. ISSN 0028-0836. 535(7610), p. 173–177. doi: 10.1038/nature18317. Full text in Research Archive

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• Canals, Mateu Montserrat; Sekulic, Nikolina & Krengel, Ute (2024). UiO structural biology core facilities. Full text in Research Archive
• Sekulic, Nikolina & Ahmad, Ahmad Ali (2023). Cryo-EM insights into the structure of the centromeric chromatin. Full text in Research Archive
• Sekulic, Nikolina; Teigen, Kelly Yong Kreutzmann; Strømme, Alexandra Nathalie & Wøien, Stine Malene Hansen (2023). Praktisk arbeid på utplassering. [Internet]. https://skolesamarbeid.oslocancercluster.no/. Full text in Research Archive
• Sekulic, Nikolina; Krengel, Ute & Brech, Andreas (2023). Minisymposium on CryoEM and CryoET: From knowledge to cures. Full text in Research Archive
• Sekulic, Nikolina (2023). Hydrogen-deuterium exchange (HDX) for monitoring protein interactions and conformational changes. Full text in Research Archive
• Sekulic, Nikolina & Segura-Pena, Dario (2022). Hydrogen-deuterium exchange (HDX) coupled to mass spectrometry for analysis of protein conformations and protein-ligand interactions. Full text in Research Archive
• Sekulic, Nikolina & Segura-Pena, Dario (2022). Hydrogen-deuterium exchange (HDX) coupled to mass spectrometry for analysis of protein conformations and protein-ligand interactions. Full text in Research Archive
• Sekulic, Nikolina; Segura-Pena, Dario; Hovet, Oda; Dawicki-McKenna, Jennine; Wøien, Stine Malene Hansen & Black, Ben E. [Show all 7 contributors for this article] (2022). Aurora B activation requires synchronized internal motion initiated by IN-box phosphorylation. Full text in Research Archive
• Segura-Pena, Dario; Hovet, Oda; Dawicki-McKenna, Jennine; Wøien, Stine Malene Hansen; Black, Ben E. & Cascella, Michele [Show all 7 contributors for this article] (2022). The structural basis of the multi-step allosteric activation of Aurora B kinase. Full text in Research Archive
• Segura-Pena, Dario; Hovet, Oda; Dawicki-McKenna, Jennine; Wøien, Stine Malene Hansen; Black, Ben E. & Cascella, Michele [Show all 7 contributors for this article] (2022). HDX and MD as tools to elucidate Aurora B kinase activation mechanism. Full text in Research Archive
• Segura-Pena, Dario; Hovet, Oda; Dawicki-McKenna, Jennine; Wøien, Stine Malene Hansen; Black, Ben E. & Cascella, Michele [Show all 7 contributors for this article] (2022). Phosphorylation structurally organizes the [Aurora B/IN-box] complex and introduces a synchronised internal motion of the enzyme resulting in activation. Full text in Research Archive
• Segura-Pena, Dario; Hovet, Oda; Dawicki-McKenna, Jennine; Wøien, Stine Malene Hansen; Black, Ben E. & Cascella, Michele [Show all 7 contributors for this article] (2022). The structural basis of the multi-step allosteric activation of Aurora B kinase. Full text in Research Archive
• Segura-Pena, Dario; Hovet, Oda; Dawicki-McKenna, Jennine; Wøien, Stine Malene Hansen; Black, Ben E. & Cascella, Michele [Show all 7 contributors for this article] (2022). The structural basis of the multi-step allosteric activation of Aurora B kinase. Full text in Research Archive
• Sekulic, Nikolina; Repasky, Gretchen & Segura-Pena, Dario (2022). Uncovering protein dynamics with hydrogen/deuterium exchange mass spectrometry in the Nordics. [Internet]. https://projects.au.dk/nordic-embl-partnership/show-news/art. Full text in Research Archive
• Sekulic, Nikolina; Yuen, Karen Wing Yee; Kelly, Alexander & Meraldi, Patrick (2022). Editor for research topic: Dynamic kinetochore. Frontiers in Cell and Developmental Biology. Full text in Research Archive
• Sekulic, Nikolina & Krengel, Ute (2022). PX-Oslo: Structural Biology Core Facilities. Full text in Research Archive
• Ali-Ahmad, Ahmad & Sekulic, Nikolina (2021). CryoEM structures of centromere complexes – collaboration between NCMM and cryoEM facility in Umeå. Full text in Research Archive
• Sekulic, Nikolina; Ali-Ahmad, Ahmad & Repasky, Gretchen (2021). Building Bridges: Sharing molecular medicine research technologies across Nordic borders. [Internet]. https://projects.au.dk/nordic-embl-partnership/show-news/art. Full text in Research Archive
• Sekulic, Nikolina & Segura-Pena, Dario (2021). Hydrogen-deuterium exchange and molecular dynamics as tools for understanding enzyme mechanism of [Aurora B/IN-box]. Full text in Research Archive
• Sekulic, Nikolina (2021). Early Career Development “How to start your own lab”. Full text in Research Archive
• Cortez, Natalia Mojica; Sekulic, Nikolina & Krengel, Ute (2020). UiO Structural Biology Core Facilities: Who Are We and How Can We Help You? Full text in Research Archive
• Mørch, Anna; Sekulic, Nikolina & Ali-Ahmad, Ahmad (2023). Structural and Functional Studies of Centromeric Protein CENP-T from Bombyx mori. Universitetet i Oslo. Full text in Research Archive
• Fløgstad, Ole Magnus; Sekulic, Nikolina & Segura-Pena, Dario (2023). Optimization of mass spectrometry techniques for epitope characterization of the interaction between HER2 and trastuzumab. Universitetet i Oslo. Full text in Research Archive Show summary

  Antibodies are well established in therapeutic treatment of cancers, autoimmune disorders, and infections for a wide variety of diseases. However, the establishment of new antibody treatments is a long process relaying on initial generation of antibodies in animal models and further optimization. To avoid the time and resource demanding process in monoclonal antibody development many attempts have been made to computationally engineer antibodies with desired binding affinities and optimal biochemical properties. Recently, the prediction of antigen-antibody specificity through deep learning has been applied to a well categorized antibody-antigen system: Human epidermal growth factor receptor 2 (HER2) and the anti-HER2 monoclonal antibody (Trastuzumab) (Mason, Friedensohn et al. 2021) (Shanehsazzadeh, Bachas et al. 2023). Binding assays of the predicted variants of trastuzumab show similar or even higher binding affinity compared to the original antibody. Interestingly, some of the predicted variant antibodies with high affinity binding have a large sequence difference, compared to the original trastuzumab, in the complementarity determining region of the heavy chain 3 (CDRH3) that is major player in epitope recognition. This observation opens the possibility that the developed antibodies bind the epitope in a different way, but the experimental techniques required to explore this possibility are limited and cumbersome. In this study, we used two mass spectrometry-based epitope mapping techniques: Hydrogen/deuterium exchange coupled with mass spectrometry (HDX-MS) and epitope excision to characterize HER2-trastuzumab interaction and whether they are useful for characterizing antibody variants. We have successfully established experimental conditions for studying the complex interaction with HDX-MS, while the epitope excision method poses some challenges for the given antibody-antigen system. The results of our study are consistent with the available high-resolution structures of HER2-trastuzumab and provide the basis for future studies on the interactions between engineered antibodies and HER2.

• Mørch, Anna & Sekulic, Nikolina (2023). Structural and Functional Studies of Centromeric Protein CENP-T from Bombyx mori. Kjemisk Institutt, Universitetet i Oslo. Full text in Research Archive
• Sekulic, Nikolina; Cascella, Michele & Hovet, Oda (2020). Molecular Dynamics Studies on the Essential Mitotic Protein Kinase – Aurora B. Universitetet i Oslo. Full text in Research Archive Show summary

  Aurora B is a serine/threonine protein kinase. Its enzymatic activity is required for accurate chromosome segregation, spindle midzone stability and cell division. Aurora B kinase is always in association with the activator protein, INCENP, and full activity requires auto-phosphorylation of both, a threonine in the activation loop of Aurora B and the serines in the TSS-motif (Threonine-Serine-Serine) present on INCENP. Structural studies have revealed profound conformational changes in the enzyme complex upon auto-phosphorylation and initial hydrogen- deuterium exchange studies indicate structural dynamic changes associated with auto-phosphorylation. In this study, we have used molecular dynamics to simulate three phosphorylation states of the enzyme complex (non-phosphorylated enzyme complex, enzyme complex phosphorylated only in the activation loop of Aurora B and enzyme complex phosphorylated in both loop of Aurora B and INCENP). We found that phosphorylation has a profound impact on the structure and dynamics of Aurora B/INCENP-complex inducing allosteric communication between INCENP and Aurora B. Upon phosphorylation of the threonine in the activation loop of Aurora B, INCENP adopts a dramatic conformational change and comes into close contact with the activation loop region. Other analyzed regions undergoing noticeable structural changes upon phosphorylation are the active site where ATP becomes more stably bound, the activation loop that becomes more rigid and the αG-helix experiencing a shift towards the core of Aurora B. Principle component analysis revealed a synchronized motion in the fully phosphorylated state of the enzyme complex with coupled open-close and activation loop motions, a feature that might contribute to increased activity. The insights into regulation of Aurora B/INCENP-complex provided in our study might help towards generating a more specific inhibitor of Aurora B in future cancer treatment.

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