Dissociation Strategies to Maximize Coverage of α‑Helical Domains in Top-Down Mass Spectrometry of Integral Membrane Proteins.
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| Title: | Dissociation Strategies to Maximize Coverage of α‑Helical Domains in Top-Down Mass Spectrometry of Integral Membrane Proteins. |
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| Authors: | Cohn, Whitaker1 (AUTHOR), Huguet, Romain2 (AUTHOR), Zabrouskov, Vlad2 (AUTHOR), Whitelegge, Julian1 (AUTHOR) jpw@chem.ucla.edu |
| Source: | Journal of the American Society for Mass Spectrometry. 6/2/2021, Vol. 32 Issue 6, p1380-1387. 8p. |
| Abstract: | Transmembrane α-helical domains of membrane proteins tend to remain structured in the gas phase, presenting a challenge for efficient electron capture/transfer dissociation during top-down dissociation mass spectrometry (MS) experiments. In this study, we compare results from different dissociation modes on a modern Orbitrap platform applied to a model integral membrane protein containing two transmembrane helices, the c-subunit of the Fo domain of the chloroplast ATP synthase. Using commercially available options, we compare collisionally activated dissociation (CAD) with the related variant higher-energy collisional dissociation (HCD) and with electron transfer dissociation (ETD). HCD performed better than CAD and ETD. A combined method utilizing both ETD and HCD (EThcD) demonstrates significant synergy over HCD or ETD alone, representing a robust option analogous to activated ion electron capture dissociation, whereby an infrared laser was used to heat the protein ion alongside electron bombardment. Ultraviolet photodissociation at 213 nm displays at least three backbone dissociation mechanisms and covered nearly 100% of backbone bonds, suggesting significant potential for this technique. [ABSTRACT FROM AUTHOR] |
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| Database: | Engineering Source |
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