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Biochemistry. 2018 Mar 13;57(10):1568-1571. doi: 10.1021/acs.biochem.8b00003. Epub 2018 Feb 23.

Sequential Protein Expression and Capsid Assembly in Cell: Toward the Study of Multiprotein Viral Capsids Using Solid-State Nuclear Magnetic Resonance Techniques.

Author information

1
Department of Chemistry and Biochemistry , The City College of New York , 160 Convent Avenue , New York , New York 10031 , United States.
2
New York Structural Biology Center , 89 Convent Avenue , New York , New York 10027 , United States.
3
PhD Programs in Biochemistry and Chemistry , The Graduate Center, City University of New York , New York , New York 10016 , United States.
4
PhD Program in Physics , The Graduate Center, City University of New York , New York , New York 10016 , United States.

Abstract

While solid-state nuclear magnetic resonance (ssNMR) has emerged as a powerful technique for studying viral capsids, current studies are limited to capsids formed from single proteins or single polyproteins. The ability to selectively label individual protein components within multiprotein viral capsids and the resulting spectral simplification will facilitate the extension of ssNMR techniques to complex viruses. In vitro capsid assembly by combining individually purified, labeled, and unlabeled components in NMR quantities is not a viable option for most viruses. To overcome this barrier, we present a method that utilizes sequential protein expression and in cell assembly of component-specifically labeled viral capsids in amounts suitable for NMR studies. We apply this approach to purify capsids of bacteriophage ϕ6 isotopically labeled on only one of its four constituent protein components, the NTPase P4. Using P4-labeled ϕ6 capsids and the sensitivity enhancement provided by dynamic nuclear polarization, we illustrate the utility of this method to enable ssNMR studies of complex viruses.

PMID:
29465229
DOI:
10.1021/acs.biochem.8b00003
[Indexed for MEDLINE]

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