TY - JOUR
T1 - Examining the Heterogeneous Genome Content of Multipartite Viruses BMV and CCMV by Native Mass Spectrometry
AU - van de Waterbeemd, Michiel
AU - Snijder, Joost
AU - Tsvetkova, Irina B.
AU - Dragnea, Bogdan G.
AU - Cornelissen, Jeroen J.
AU - Heck, Albert J.R.
PY - 2016
Y1 - 2016
N2 - Since the concept was first introduced by Brian Chait and co-workers in 1991, mass spectrometry of proteins and protein complexes under non-denaturing conditions (native MS) has strongly developed, through parallel advances in instrumentation, sample preparation, and data analysis tools. However, the success rate of native MS analysis, particularly in heterogeneous mega-Dalton (MDa) protein complexes, still strongly depends on careful instrument modification. Here, we further explore these boundaries in native mass spectrometry, analyzing two related endogenous multipartite viruses: the Brome Mosaic Virus (BMV) and the Cowpea Chlorotic Mottle Virus (CCMV). Both CCMV and BMV are approximately 4.6 megadalton (MDa) in mass, of which approximately 1 MDA originates from the genomic content of the virion. Both viruses are produced as mixtures of three particles carrying different segments of the genome, varying by approximately 0.1 MDA in mass (~2%). This mixture of particles poses a challenging analytical problem for high-resolution native MS analysis, given the large mass scales involved. We attempt to unravel the particle heterogeneity using both Q-TOF and Orbitrap mass spectrometers extensively modified for analysis of very large assemblies. We show that manipulation of the charging behavior can provide assistance in assigning the correct charge states. Despite their challenging size and heterogeneity, we obtained native mass spectra with resolved series of charge states for both BMV and CCMV, demonstrating that native MS of endogenous multipartite virions is feasible. [Figure not available: see fulltext.]
AB - Since the concept was first introduced by Brian Chait and co-workers in 1991, mass spectrometry of proteins and protein complexes under non-denaturing conditions (native MS) has strongly developed, through parallel advances in instrumentation, sample preparation, and data analysis tools. However, the success rate of native MS analysis, particularly in heterogeneous mega-Dalton (MDa) protein complexes, still strongly depends on careful instrument modification. Here, we further explore these boundaries in native mass spectrometry, analyzing two related endogenous multipartite viruses: the Brome Mosaic Virus (BMV) and the Cowpea Chlorotic Mottle Virus (CCMV). Both CCMV and BMV are approximately 4.6 megadalton (MDa) in mass, of which approximately 1 MDA originates from the genomic content of the virion. Both viruses are produced as mixtures of three particles carrying different segments of the genome, varying by approximately 0.1 MDA in mass (~2%). This mixture of particles poses a challenging analytical problem for high-resolution native MS analysis, given the large mass scales involved. We attempt to unravel the particle heterogeneity using both Q-TOF and Orbitrap mass spectrometers extensively modified for analysis of very large assemblies. We show that manipulation of the charging behavior can provide assistance in assigning the correct charge states. Despite their challenging size and heterogeneity, we obtained native mass spectra with resolved series of charge states for both BMV and CCMV, demonstrating that native MS of endogenous multipartite virions is feasible. [Figure not available: see fulltext.]
KW - Brome Mosaic Virus (BMV)
KW - Cowpea Chlorotic Mottle Virus (CCMV)
KW - Genome
KW - Native mass spectrometry
KW - Orbitrap
KW - Q-TOF
KW - Virus
UR - http://www.scopus.com/inward/record.url?scp=84971233907&partnerID=8YFLogxK
U2 - 10.1007/s13361-016-1348-6
DO - 10.1007/s13361-016-1348-6
M3 - Article
AN - SCOPUS:84971233907
SN - 1044-0305
VL - 27
SP - 1000
EP - 1009
JO - Journal of the American Society for Mass Spectrometry
JF - Journal of the American Society for Mass Spectrometry
IS - 6
ER -