Project

Discipline

Protein Engineering; Computational Structural Biology; Protein Design; Protein Recognition; X-ray crystallography; Molecular Dynamics; NMR; Protein Structure

Lead
Lay summary
The future study of cellular regulation, epigenetics and disease states will require detection tools for proteins that are as versatile as oligonucleotides are for DNA. Today, the fundamental difference between the detection of proteins and that of DNA is that, for each protein and each modification, an individual reagent has to be separately generated and validated, while for DNA, synthetic oligonucleotides give access to a completely generic detection technology, using only four building blocks, known complementarity rules and automated chemical synthesis. The grand ambition of this Sinergia project is to create a similar, completely modular detection technology for polypeptides as well. This will require that a peptide (e.g., from an unstructured region of a protein, a denatured form of the protein or proteolytic digest) bind to a modular counterpart. Armadillo repeat proteins provide the basis for such a binding mode of a peptide.

Direct link to Lay Summary

Last update: 21.02.2013

Active participation

Number	Title	Start	Funding scheme

The future study of cellular regulation, epigenetics and disease states will require detection tools for proteins that are as versatile as oligonucleotides are for DNA. Today, the fundamental difference between the detection of proteins and that of DNA is that, for each protein and each modification, an individual reagent has to be separately generated and validated, while for DNA, synthetic oligonucleotides give access to a completely generic detection technology, using only four building blocks, known complementarity rules and automated chemical synthesis. The grand ambition of this Sinergia project is to create a similar, completely modular detection technology for polypeptides as well. This will require that a peptide (e.g., from an unstructured region of a protein, a denatured form of the protein or proteolytic digest) bind to a modular counterpart. Armadillo repeat proteins provide the basis for such a binding mode of a peptide. Over the first term of this grant, four strong teams of scientists, all located at the University of Zurich, closely collaborated bringing together complementary expertise in protein engineering to initiate this project: evolutionary engineering, NMR, X-ray crystallography and structure-based computation. This unique interdisciplinary collaboration has resulted in remarkable progress in this challenging endeavor, (i) having allowed to determine several crystal structures of consensus Armadillo repeat proteins with important feed-back for further design, (ii) solved many of the NMR assignment problems of these challenging repeat proteins, (iii) introduced many essential design proposals through computation and (iv) verified these proposals experimentally, including selecting a first peptide binder from a designed library.This project will be continued, exploiting the technical progress that has been made. Through the engineering of highly "crystallizable" fusion proteins, a more rapid feedback from the effect of mutations on the structure and that of the bound peptide will be obtained. Through the completion of the NMR assignment, rapid information on the location of the bound peptide will be obtained. Computation will be crucial to test ideas for modifications in the structure or of the binding pockets. Great progress in setting up high-throughput display technologies and novel systems will allow an experimental approach to the fine-tuning of the building blocks.We believe that this project, once completed, may have significant effect on future research in cell biology. Regarding first applications, we will focus our efforts on posttranslational modifications, such as site-specific and protein-specific detection of phosphorylation and histone modifications.As has been shown in the last phase, the participating students and postdoctoral fellows, which take active parts in very frequent common progress meetings, are exposed to first-hand accounts of a wide range of different state-of-the-art technologies relevant in modern protein science, and have thus greatly profited from this interdisciplinary research approach to protein engineering.