کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
5131372 | 1490887 | 2016 | 8 صفحه PDF | دانلود رایگان |
- CE-ESI-MS is proposed for protein conformational analysis.
- The amyloidogenic protein beta2-microglobulin is used as a model.
- Protein conformers are studied under native and misfolding conditions.
- New sheathless interfacing is best suited to preserve protein structure integrity.
- CE-ESI-TOF MS reliably assigns protein forms that differ by 1Â Da.
In this work we explored the feasibility of different CE-ESI-MS set-ups for the analysis of conformational states of an intact protein. By using the same background electrolyte at quasi physiological conditions (50Â mM ammonium bicarbonate, pH 7.4) a sequential optimization was carried out, initially by evaluating a sheath-liquid interface with both a single quadrupole (SQ) and a time-of-flight (TOF) mass spectrometer; then a sheathless interface coupled with high-resolution QTOF MS was considered. Beta2-microglobulin has been taken as a model, as it is an amyloidogenic protein and its conformational changes are strictly connected to the onset of a disease. The separation of two conformers at dynamic equilibrium is achieved all the way down to the MS detection. Notably, the equilibrium ratio of the protein conformers is maintained in the electrospray source after CE separation. Strengths and weaknesses of each optimized set-up are emphasized and their feasibility in unfolding studies is evaluated. In particular, ESI-TOF MS can assign protein forms that differ by 1Â Da only and sheathless interfacing is best suited to preserve protein structure integrity. This demonstrates the CE-ESI-MS performance in terms of separation, detection and characterization of conformational species that co-populate a protein solution.
Journal: Analytica Chimica Acta - Volume 945, 16 November 2016, Pages 102-109