2019 Fiscal Year Final Research Report
Enabling NMR studies of sparsely labelled large proteins by automated assignment
| Project/Area Number |
17K07312
|
|---|
| Research Category |
Grant-in-Aid for Scientific Research (C)
|
| Allocation Type | Multi-year Fund |
|---|
| Section | 一般 |
|---|
| Research Field |
Structural biochemistry
|
|---|
| Research Institution | Tokyo Metropolitan University |
|---|
Principal Investigator |
Guentert Peter 首都大学東京, 理学研究科, 客員教授 (20392110)
|
|---|
| Co-Investigator(Kenkyū-buntansha) |
池谷 鉄兵 首都大学東京, 理学研究科, 助教 (30457840)
|
|---|
| Project Period (FY) |
2017-04-01 – 2020-03-31
|
| Keywords | protein NMR / in-cell NMR / resonance assignment / automated assignment / methyl groups / isotope labeling |
|---|
| Outline of Final Research Achievements |
Proteins that are large, membrane-bound, or studied in living cells by in-cell NMR can in general not be assigned by the conventional solution NMR method that relies on uniform 13C/15N-labeling because the resonance lines become too broad and overlapping. Interpretable spectra can be restored by sparse labeling of methyl groups although resonance assignments remain difficult to obtain. Here we developed the MethylFLYA method that can assign large, methyl-labeled proteins using NOESY spectra in conjunction with a known 3D structure. MethylFLYA finds assignments by optimizing a mapping between expected peaks based on the protein sequence, and the measured peaks identified by peak picking. The new approach has been applied to large proteins up to 468 kDa size and to proteins in living cells.
|
| Free Research Field |
Biomolecular NMR spectroscopy
|
| Academic Significance and Societal Importance of the Research Achievements |
The methods developed in this research project makes new classes of proteins more easily accessible to detailed NMR studies. Previously, NMR resonance assignments for these proteins could only be determined by time-consuming experimental methods such as extensive mutagenesis.
|
