SINGLEMEMB · Real-time tracking of single membrane translocases
7РП — „Хора“ (Действия „Мария Кюри“)
- Период
- 2013-08-01 → 2017-07-31
- Финансиране от ЕС
- 100 000 €
- Участници
- 1
- Схема
- MC-CIG
Линиите свързват координатора с партньорите.
Накратко на български
Протеиновите транслокази, които пренасят молекули през клетъчните мембрани, се проследяват в реално време чрез специални оптични инструменти. Това помага за по-доброто разбиране на процесите по транспортиране и разграждане на протеините в клетката.
Кратко обяснение, генерирано от езиков модел по текста на CORDIS. Оригиналът е по-долу.
Резултати накратко
Real-time tracking of single membrane translocases
The threading of proteins through narrow channels within a cell is a crucial biological process, which is essential for cellular protein trafficking and protein degradation. The motor proteins driving this translocation process are called protein translocases. The Aubin-Tam group is developing biophysical tools to track the dynamics of protein translocation at the single-molecule level. The project objectives since the beginning of the Marie Curie CIG consisted of integrating a lipid bilayer electrophysiology within an optical tweezers system and of integrating a membrane translocase into the optical tweezers assay. During the second period of the Marie Curie CIG, free-standing phospholipid bilayers were interfaced with optical tweezers, with access to both lipid bilayer leaflets. FtsH has also been further structurally and biochemically characterized. Since the beginning of the project, a strategy has been developed to functionalize styrene maleic acid lipid nanodiscs with small molecules. Labeling with a biotin molecule and with a fluorophore has been demonstrated. In parallel, a new type of microdevice has been designed and fabricated to form stable free-standing phospholipid bilayers. One major difficulty was to optically trap in close vicinity of the lipid membrane. A quantity of solvent, known as solvent annulus, is usually retained between the two leaflets at the edge of the membrane. Unfortunately, this annulus creates optical aberrations preventing optical trapping close to the membrane. To overcome this issue, the Aubin-Tam group identified a right combination of solvent and material for the flow cell, which led to successful optical trapping in close vicinity of the membrane. The tools developed in this project could advance single-molecule investigations of membrane proteins, which represent 30% of all proteins and are prime drug targets. They will also find application in the study of the wide range of mechanical processes which occur at the cell membrane, such as lipid nanotube formation, flagellar locomotion, protein translocation or endocytosis. https://sites.google.com/site/aubintamgroup/
Текст от CORDIS, на английски · Данни: CORDIS, © Европейски съюз
Цел на проекта
The threading of proteins through narrow channels within a cell is a crucial biological process, which is essential for cellular protein trafficking and protein degradation. This translocation process is also used by pathogens such as anthrax and C. Difficile, which inject their lethal toxins through the cell membrane in order to hijack vital cell machinery. The motor proteins driving this translocation process are called protein translocases. Despite the ubiquity of protein translocation in biology, the underlying mechanisms remain poorly understood. Improving our knowledge of translocation will have implications on our understanding of severe pathologies, which have been linked to the dysfunction of translocases (e.g., cancers, Alzheimer’s).I intend to design novel biorelevant in vitro tools to investigate the inner workings of membrane-associated protein translocases. The translocase will be reconstituted in a cell-like microenvironment, while allowing real time control over biologically appropriate external conditions that may affect its function in vivo (e.g., voltage/pH gradient across membrane). Our innovative approach is based on the integration of novel lipid systems into a combination of optical tweezers and electrophysiology techniques. The first translocase targeted is FtsH, whose role is to dislocate proteins from membranes. We will directly measure the translocation velocity and step size of this membrane-associated translocase with unprecedented detail, while being able to control the protonmotive force.These new assays will advance considerably single-molecule investigations of membrane proteins, which represent 30% of all proteins and are prime drug targets. This work will lay foundations towards elucidating the mechanisms behind other crucial membrane translocases, in particular transmembrane protein transporters.
Оригинален текст от CORDIS (на английски).
Участници
- TECHNISCHE UNIVERSITEIT DELFT · DelftКоординаторНидерландия
Връзки
Данни: CORDIS, © Европейски съюз