  Mass spectrometry (MS) is used widely in biomolecular structural analysis and
is particularly dominant in the study of proteins. Despite its considerable
power, state-of-the-art protein MS frequently suffers from limited reliability
of spectrum-to-structure assignments. This could not be solved fully by the
dramatic increase in mass accuracy and resolution of modern MS instrumentation
or by the introduction of new fragmentation methods. Here we present a new kind
of two-dimensional mass spectrometry for high fidelity determination of a
biomolecular primary structure based on partial covariance mapping. Partial
covariance two-dimensional mass spectrometry (pC-2DMS) detects intrinsic
statistical correlations between biomolecular fragments originating from the
same or consecutive decomposition events. This enables identification of pairs
of ions produced along the same fragmentation pathway of a biomolecule across
its entire fragment mass spectrum. We demonstrate that the fragment-fragment
correlations revealed by pC-2DMS provide much more specific information on the
amino acid sequence and its covalent modifications than the individual fragment
mass-to-charge ratios on which standard one-dimensional MS is based. We
illustrate the power of pC-2DMS by using it to resolve structural isomers of
combinatorially modified histone peptides inaccessible to standard MS.
