H2020Индивидуална стипендия2015–2017

FUNC NGS · Deep screening of proteins with a next generation sequencing platform

„Хоризонт 2020“ — Действия „Мария Склодовска-Кюри“

Период: 2015-08-01 → 2017-07-31
Финансиране от ЕС: 195 455 €
Участници: 1
Схема: MSCA-IF-EF-ST

Линиите свързват координатора с партньорите.

Накратко на български

Методът за анализ на протеинови взаимодействия чрез секвениране от ново поколение позволява бързото тестване на милиони варианти на протеини. Това помага за подобряване на ефективността на лекарствени средства, базирани на протеини.

Този кратък обзор е генериран от изкуствен интелект

Кратко обяснение, генерирано от езиков модел по текста на CORDIS. Оригиналът е по-долу.

Резултати накратко

Deep screening of proteins with a next generation sequencing platform

Protein engineering is often required to improve the efficiency of protein-based therapeutic agents. Despite the remarkable advancements in computer-aided protein design, the most potent candidates for a binding interaction are still found by experimental methods. The analysis of more than a few hundred to thousand individual variants requires automated high-throughput screening platforms, but the throughput of even the most modern microtiter plate screening methods is still a tiny fraction of the size of combinatorial repertoires operated with phage and ribosome display (>109). Therefore, better assays for quantitative protein interaction analysis are still highly demanded. Next generation sequencing (NGS) has become an indispensable tool in visualizing the effect of mutations in proteins on a global scale using hundreds of millions of dna sequences. Several chemistries have been developed for NGS, of which, the most used today are Ion Torrent semiconductor sequencing technology, Roche 454 pyrosequencing and Illumina sequencing by synthesis technology. The objective of this proposal was to set up a generalized method for analysing protein interactions using a cell-free display technique combined to a next-generation sequencing platform in order to increase the throughput of protein interaction analysis. A method was designed that used microbeads and protein DNA conjugates as the media for protein reactions. The technology was aimed to be compatible with most next generation sequencing platforms for analysis. Research towards the final deep screening technology was supported by other technology development projects targeting different aspects of bead surface display methods.

Текст от CORDIS, на английски · Данни: CORDIS, © Европейски съюз

Цел на проекта

Next generation sequencing (NGS) has become an indispensable tool for system biology and has recently been adopted in protein engineering science as well. This proposal describes the development of a highly generalizable technique, termed FUNC-NGS, which links in situ functional information of protein activity with the function-determining DNA sequence in a massively parallel manner opening exciting possibilities, e.g., in drug discovery. In this method a cell-free display platform is combined with NGS by modifying protein display units to contain unique homing oligonucleotide sequences. These units are subsequently targeted to bind monoclonal DNA clusters in the flow cell of an Illumina sequencer by complementary DNA hybridization. FUNC-NGS is readily compatible with a paired-end sequencing programme on the Illumina platform and the number of screening units is only limited by the number of DNA clusters on the flow cell. As the first application challenge, a new variant of a modified bacterial adhesin, Spycatcher, will be created with an altered specificity for synthetic biology applications. Novel Spycatcher-SpyTag pairs are intensively desired but their creation is demanding. Solving this problem will be a perfect showcase for this radically novel deep screening technology. At the mature stage FUNC-NGS will be a transformative platform in therapeutic protein development, functional genomics and personalized medicine. The proposed work will be carried out at Dr. Hollfelder´s laboratory at the University of Cambridge with a short secondment at Medimmune. Hollfelder group has developed SNAP display, which is a FUNC-NGS-compatible in vitro protein display system. Furthermore, the University hosts excellent facilities for NGS sequencing and experienced bioinformatics support is available. The planned research programme synergistically combines my expertise in molecular biology method development with the hosts expertise in cell-free display and IVC-techniques.

Оригинален текст от CORDIS (на английски).

Участници

THE CHANCELLOR MASTERS AND SCHOLARS OF THE UNIVERSITY OF CAMBRIDGE · CAMBRIDGEКоординаторОбединеното кралство