Nucleic Acids Research, 2009, Vol. 37, No. 14 4551 Figure 3. Analysis of CrgA and CrgA–DNA complexes by non-denaturing mass spectrometry. The predominant charge state series in the nano- electrospray mass spectrum shown in (A) corresponds to the +36 to +43 charge states of CrgA (3 mM) with an octameric stoichiometry. A small amount of hexameric CrgA was also observed (indicated by black circles) and the insert at the top of the panel conﬁrms there are no other stoichiometries over a wide m/z range. (B) The CrgA octamer under activating MS conditions, resulting in the formation of a ‘stripped’ heptameric species (insert). (C) The mixed population of CrgA–DNA complexes observed by MS, these include CrgA8DNA1, CrgA8DNA2, CrgA8DNA3 and CrgA16DNA4. A spectrum of DNA (80 mM) alone is shown as an insert to (C). In this insert four charge state series were identiﬁed with charges states from +10 to +14. The full length DNA corresponds to series D, while the smaller masses (series A–C) likely correspond to various nucleic acid truncations of the DNA. Figure 4. Comparison of CrgA and CbnR. (A) Dimeric unit of CrgA. (B) Dimeric unit of CbnR (PDB code 1IZ1) and CbnR tetramer (Insert). preventing a comparable interface forming in this protein (Figure 4B). CrgA is atypical since 14/15 closest structural neighbors identiﬁed using a Dali search contain a helix at this position. To probe the relationship between CrgA and other members of the LTTR family we generated a cluster map of 4843 LTTR sequences with the program CLANS which uses pair-wise BLAST similarity scores (47).