Project 10: Mining the functional microproteome landscape using thermal proteome profiling

Only a fraction of the genome codes for the proteins in standard reference catalogs. Many small open reading frames (smORFs) scattered across the genome are translated into microproteins that these catalogs miss, and while growing numbers of microproteins are being discovered, their biological functions and the molecular interactions through which they act remain largely unknown. Thermal proteome profiling (TPP), a mass-spectrometry method that measures how the thermal stability of a protein shifts when its binding partners or conditions change, offers a way to assign function to these previously hidden proteins.

This project will systematically characterize the functional landscape of microproteins in the enteropathogen Salmonella enterica serovar Typhimurium, using TPP across smORF deletion strains and multiple conditions to build a comprehensive interaction map for the Salmonella microproteome. It sits at the center of ORFeus Work Package 2, which moves from discovery to function: the structural and biophysical data it generates feed an iterative design-test cycle with a computational protein-design project, while the standardized TPP workflows developed here are adapted for host-pathogen proteomics elsewhere in the network. The work also connects to evolutionary analysis of microprotein function and to the wider consortium through the shared ORFeome platform.

Main tasks

• Develop and standardize thermal proteome profiling protocols to measure changes in protein stability and abundance across a panel of smORF deletion strains and multiple growth conditions in Salmonella.
• Integrate the thermal proteome profiling data with protein structure prediction (for example AlphaFold and RoseTTAFold), working with the computational protein design project of another doctoral candidate (DC) in the network, here DC6, to infer protein-protein interactions involving microproteins.
• Validate high-confidence interactions in vitro using biophysical methods such as biolayer interferometry and SEC-MALS, building a validated interaction map for the Salmonella microproteome.
• Collaborate across the ORFeus network by sharing standardized TPP workflows with related projects, including host-pathogen proteomics in DC4, contribute datasets to the shared ORFeome platform, and producing the project’s scientific report.

Methods and platforms: thermal proteome profiling (TPP) and quantitative mass spectrometry-based proteomics, construction and phenotyping of smORF deletion strains in Salmonella, protein structure prediction (AlphaFold, RoseTTAFold) for interaction modeling, and in vitro biophysical validation (biolayer interferometry, SEC-MALS), with datasets shared through the ORFeome platform.

Secondment: you will spend around three months at Bruker (Bremen, Germany), optimizing the thermal proteome profiling workflows on advanced mass-spectrometry platforms. You will also be guided by an independent academic advisor, with the possibility of a short, primarily virtual research exchange to strengthen the project.

Your profile

MSCA eligibility

You must meet all of the following on your recruitment date:

• You do not already hold a doctoral degree. If you have defended a doctoral thesis but the degree has not yet been formally awarded, you are not eligible.
• Mobility rule: you must not have lived or carried out your main activity (work, studies, and so on) in Belgium for more than 12 months in the 36 months immediately before your recruitment date. Compulsory national service, short stays such as holidays, and time spent in a procedure to obtain refugee status under the Geneva Convention do not count toward the 12 months.
• You hold, or will hold before the start date, a degree that formally entitles you to enroll in a doctorate, and you can enroll in the doctoral program at Ghent University.
• Candidates of any nationality may apply. There is no limit on prior research experience, as long as you do not already hold a doctorate.

Project-specific profile

• A master's degree (or equivalent) in biochemistry, molecular biology, microbiology, biotechnology, biomedical sciences, analytical chemistry, or a related life-science field.
• Hands-on laboratory experience, ideally including protein biochemistry, mass spectrometry, or proteomics, and comfort working with quantitative data.
• Desirable: experience with mass-spectrometry-based proteomics, bacterial genetics or Salmonella work, biophysical interaction methods (for example biolayer interferometry or SEC-MALS), and an interest in microproteins and the dark proteome.
• Good written and spoken English.
• Motivation for interdisciplinary, collaborative research, and willingness to travel for the secondment and network events.

What we offer

We offer a full-time position