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Molecular Neurobiology

Department                             Bionanoscience

Principal investigator          Dimphna Meijer

E-mail address                      d.h.m.meijer@tudelft.nl

Website                                  www.dimphnameijerlab.org

 

Engineering Novel Cell-Adhesion Receptors Using the PURE System in Synthetic Cells

Suitable as a BEP? No

Suitable as a MEP? Yes

Suitable as an Academic Research Project? No

Techniques:
  • Protein expression in both eukaryotic and prokaryotic cell lines
  • Purification using the ÄKTA Pure system
  • Protein engineering performed through molecular cloning techniques and in-silico design tools

Synthetic cells are defined as engineered vesicles, capable of mimicking biological activities. They are emerging as potential tools for applications across various fields, particularly in the medical and pharmaceutical sectors, but also in environmental and energy-related areas.
This project aims to investigate the sensing and interaction capabilities of synthetic cells mediated by cell adhesion receptors, as well as the intracellular mechanisms involved in sensing. The focus will be on the design of Engineered Novel Cell-Adhesion Receptors that enable these systems to adapt to their surrounding environment. The student will contribute to the design and expression of novel cell-adhesion receptors in synthetic cells, with the goal of characterizing their structural and biophysical properties.

Further reading (click to link to article)

Sato W, Zajkowski T, Moser F, Adamala KP. Synthetic cells in biomedical applications. Wiley Interdiscip Rev Nanomed Nanobiotechnol. 2022 Mar;14(2):e1761. doi: 10.1002/wnan.1761. Epub 2021 Nov 1. PMID: 34725945; PMCID: PMC8918002. Kozlova I, Sah S, Keable R, Leshchyns’ka I, Janitz M, Sytnyk V. Cell Adhesion Molecules and Protein Synthesis Regulation in Neurons. Front Mol Neurosci. 2020 Nov 12;13:592126. doi: 10.3389/fnmol.2020.592126. PMID: 33281551; PMCID: PMC7689008.

Molecular and structural characterization of candidate biomarker NPTX in frontotemporal dementia

Suitable as a BEP? No

Suitable as a MEP? Yes

Suitable as an Academic Research Project? No

Techniques:
  • Protein expression in mammalian cells and purification with the AKTA pure system.
  • Techniques for characterization such as SEC-MALS
  • Mass Photometry and thermostability assays.
  • Negative Staining
  • Cryo EM

Neuronal pentraxins (NPTXs) have emerged as promising biomarkers for neurodegenerative diseases. In collaboration with the Neurology Department of the Erasmus Medical Center (EMC), which specializes in Frontotemporal Dementia (FTD), this project aims to solve the molecular and structural characterization of NPTXs and to identify novel binding partners of NPTXs in human brain tissue.
The master student will characterize the neuronal pentraxins (NPTX1, NPTX2, and NPTXR) at both the molecular and biophysical levels. This will involve studying their structural and functional properties and investigating complex formation between these proteins and potential interaction partners. Through this work, the student will contribute to a better understanding of the molecular mechanisms underlying neuronal pentraxin function and their potential role in FTD.

Further reading (click to link to article)

1. de San Jose, N.G., et al (2022). Neuronal pentraxins as biomarker of synaptic activity: from physiological functions to pathological changes in neurodegeneration. Journal of Neural Transmission 129, 207-230. 2. Xu, D., et al (2003). Narp and NP1 form heterocomplexes that function in developmental and activity-dependent synaptic plasticity. Neuron 39, 513-528.

Unravelling the impact of patient CAMK2G mutations on protein activity and neuronal function

Suitable as a BEP? Yes

Suitable as a MEP? Yes

Suitable as an Academic Research Project? Yes

Techniques:
  • Mammalian cell culture
  • Protein purification
  • Western blot
  • Electron microscopy/Cryo-EM
  • Molecular biology

Synaptic plasticity, a fundamental process in brain function, underpins the remarkable adaptability of the nervous system to external stimuli. In this process, the calcium/calmodulin-dependent protein kinase II (CAMK2) family emerges as a pivotal player. The CAMK2 family is a Ser/Thr protein kinase family consisting of four isozymes (CAMK2A, CAMK2B, CAMK2G, and CAMK2D). Recently, multiple de novo mutations in CAMK2G gene were identified among patients suffering from intellectual disability, highlighting the importance of CAMK2G in the brain. However, the mechanisms through which these mutations contribute to the onset of the disease remain poorly characterized. In this project we will use a combination of structural biology and biophysical assays to investigate the effects of these mutations on CAMK2G’s structure and activity. This project is a collaborative project between TU Delft and Erasmus MC.

Further reading (click to link to article)

Proietti Onori M, Koopal B, Everman DB, et al. The intellectual disability-associated CAMK2G p.Arg292Pro mutation acts as a pathogenic gain-of-function. Human Mutation. 2018; 39: 2008–2024. Myers, J., Zaegel, V., Coultrap, S. et al. The CaMKII holoenzyme structure in activation-competent conformations. Nat Commun 8, 15742 (2017). https://pubmed.ncbi.nlm.nih.gov/30184290/

(Example) projects submitted by lab in past years

(2024-2025) Structural characterisation of FAM171A2 – FAM171B protein complex

Supervisor: Leanid Kresik, L.K.Kresik@tudelft.nl

In this project, we plan to perform structural biology studies of neuronal transmembrane proteins FAM171A2 and FAM171B from the FAM171 protein family. FAM171A2 and FAM171B are single-pass transmembrane proteins expressed in the brain interacting with each other. The existing literature provides limited information about the localization and functions of these proteins, and there is currently no available data about their structure. Using mammalian protein expression systems and protein purification techniques we will produce and purify single proteins and reconstitute FAM171-A2B complex in vitro. Next, we will utilise advanced electron microscopy technics such as negative stain electron microscopy and cryogenic electron microscopy for 3D molecular structure determination of the protein complex. Structural data will be complemented with other biophysical assays, such as Mass Photometry and Size-Exclusion Chromatography combined with Multi-Angle Light Scattering.

Techniques

  • Cell culture
  • Protein purification
  • Size-Exclusion Chromatography
  • Negative stain electron microscopy
  • Cryogenic electron microscopy
  • Mass Photometry

 

(2024-2025) Unravelling the impact of patient CAMK2G mutations on protein activity and neuronal function

Supervisor: Angelica Casotto, a.c.casotto@tudelft.nl

Synaptic plasticity, a fundamental process in brain function, underpins the remarkable adaptability of the nervous system to external stimuli. In this process, the calcium/calmodulin-dependent protein kinase II (CAMK2) family emerges as a pivotal player. The CAMK2 family is a Ser/Thr protein kinase family consisting of four isozymes (CAMK2A, CAMK2B, CAMK2G, and CAMK2D). Recently, multiple de novo mutations in CAMK2G gene were identified among patients suffering from intellectual disability, highlighting the importance of CAMK2G in the brain. However, the mechanisms through which these mutations contribute to the onset of the disease remain poorly characterized. In this project we will use a combination of structural biology and biophysical assays to investigate the effects of these mutations on CAMK2G’s structure and activity.

Techniques

  • Mammalian cell culture
  • Protein purification
  • Western blot
  • Electron microscopy
  • Molecular biology