Quantifying the heterogeneity of macromolecular machines by mass photometry.
Nat Commun
; 11(1): 1772, 2020 04 14.
Article
in En
| MEDLINE
| ID: mdl-32286308
ABSTRACT
Sample purity is central to in vitro studies of protein function and regulation, and to the efficiency and success of structural studies using techniques such as x-ray crystallography and cryo-electron microscopy (cryo-EM). Here, we show that mass photometry (MP) can accurately characterize the heterogeneity of a sample using minimal material with high resolution within a matter of minutes. To benchmark our approach, we use negative stain electron microscopy (nsEM), a popular method for EM sample screening. We include typical workflows developed for structure determination that involve multi-step purification of a multi-subunit ubiquitin ligase and chemical cross-linking steps. When assessing the integrity and stability of large molecular complexes such as the proteasome, we detect and quantify assemblies invisible to nsEM. Our results illustrate the unique advantages of MP over current methods for rapid sample characterization, prioritization and workflow optimization.
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Mass Spectrometry
/
Cryoelectron Microscopy
Limits:
Animals
Language:
En
Journal:
Nat Commun
Journal subject:
BIOLOGIA
/
CIENCIA
Year:
2020
Document type:
Article
Affiliation country:
United kingdom