Extended Data Fig. 5: Native-MS and the corresponding particle types observed in cryo-EM of the GudB enriched preparation of the GudB-GltAB complex.
From: A counter-enzyme complex regulates glutamate metabolism in Bacillus subtilis
(a) Native-MS spectra showing different species of GudB-GltAB complex. These species primarily differ in the number of GltAB heterodimers attached to the GudB hexamer (from 3-6). Charge states (z) and the difference from theoretical mass (ΔT) is indicated for each species. While the mass of the fully assembled complex agreed well with the expected mass, high ΔT values of the other species could be because of degradation of some of the component proteins during the extended incubation step with E. coli lysate (Methods section) during the purification process. The inset shows the SDS-PAGE of GudB-GltAB complex used for the native-MS. The sample was prepared after enriching the B. subtilis lysate with recombinant GudB expressed and purified from E. coli (see Protein expression and purification, Methods). The image is from a single experiment and is a representative of at least two independent experiments. This sample contained a higher fraction of GudB and was also used for cryo-EM to obtain the high-resolution structure of GudB6-GltA2B2 complex (Fig. 5). (b) Preliminary cryo-EM maps corresponding to different particle types (GudB6-GltA2-4B2-4) observed. The key difference between particles is the number of GltAB subunits - the least being two (grey) and the maximum four (violet).
