Activation mechanisms of transcriptional regulator CooA revealed by small-angle X-ray scattering.
CooA, a heme-containing transcriptional activator, binds CO to the heme moiety and then undergoes a structural change that promotes the specific binding to the target DNA. To elucidate the activation mechanism coupled to CO binding, we investigated the CO-dependent structural transition of CooA with small-angle X-ray scattering (SAXS). In the absence of CO, the radius of gyration Rg and the second virial coefficient (A2) were 25.3(+/-0.5)A and -0.39(+/-0.25) x 10(-4)ml mol g(-2), respectively. CO binding caused a slight increase in Rg (by 0.5A) and a marked decrease in A2 (by 5.09 x 10(-4)ml mol g(-2)). The observed decrease in A2 points to higher attractive interactions between CO-bound CooA molecules in solution compared with CO-free CooA. Although the minor alternation of Rg rules out changes in the overall structure, the marked change in the surface properties points to a CO-induced conformational transition. The experimental Rg and SAXS curves of the two states did not agree with the crystal structure of CO-free CooA. We thus simulated the solution structures of CooA based on the experimental data using rigid-body refinements as well as low-resolution model reconstructions. Both results demonstrate that the hinge region connecting the N-terminal heme domain and C-terminal DNA-binding domain is kinked in CO-free CooA, so that the two domains are positioned close to each other. The CO-dependent structural change observed by SAXS corresponds to a slight swing of the DNA-binding domains away from the heme domains coupled with their rotation by about 8 degrees around the axis of 2-fold symmetry.
RIKEN Harima Institute/SPring-8, Structural Biochemistry Laboratory, 1-1-1 Kouto, Mikazuki, Sayo, Hyogo 679-5148, Japan.
Shuji
S
Akiyama
Tetsuro
T
Fujisawa
Koichiro
K
Ishimori
Isao
I
Morishima
Shigetoshi
S
Aono
D000596:Amino Acids / Q000737:chemistry; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D002248:Carbon Monoxide / Q000737:chemistry; D018360:Crystallography, X-Ray; D000242:Cyclic AMP / Q000737:chemistry; D004247:DNA / Q000737:chemistry; D015964:Gene Expression Regulation, Bacterial; D006418:Heme / Q000737:chemistry; D006420:Hemeproteins / Q000737:chemistry* / Q000378:metabolism*; D007700:Kinetics; D008027:Light; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012542:Scattering, Radiation; D013053:Spectrophotometry; D015534:Trans-Activators / Q000737:chemistry* / Q000378:metabolism*; D015533:Transcriptional Activation; D014466:Ultraviolet Rays; D014965:X-Rays
D000596:Amino Acids; D001426:Bacterial Proteins; C108776:CooA protein, Rhodospirillum rubrum; D006420:Hemeproteins; D015534:Trans-Activators; D006418:Heme; D002248:Carbon Monoxide; D004247:DNA; D000242:Cyclic AMP