NTP-binding properties of the blue-light receptor YtvA and effects of the E105L mutation.
YtvA is a blue-light-sensing protein from Bacillus subtilis related to plant phototropins. It carries a LOV (light, oxygen and voltage) domain, binding FMN (flavin mononucleotide) as chromophore, and a STAS (sulphate transporters and antisigma-factor antagonists) domain with poorly characterized function. We have recently shown that YtvA binds triphosphate nucleotides (NTP) and highlighted a structural similarity between the STAS domain and small GTP-binding proteins. In this work we further investigated the NTP-binding properties of YtvA, employing a fluorescent derivative of GTP (GTP(TR)) and mutagenesis experiments. The main results are as follows: (a) competition experiments indicate that the affinity of YtvA for GTP is much higher than that for GDP and GMP. (b) Blue-light-induced structural changes are transmitted from the LOV core to the NTP-binding cavity, establishing a possible intraprotein signal-transduction pathway. (c) A mutation in the central beta-scaffold of the LOV core, E105L, impairs the light-driven spectroscopic changes of bound GTP(TR). This result is supported by circular dichroism data, in that YtvA-E105L does not show the light-induced conformational change in the turn fraction that characterizes YtvA, implying that E105 is functionally important. (d) In the structural model of the LOV-STAS complex, based on docking algorithms, the interface includes the Ibeta-Hbeta loop on the LOV core, as well as parts of the central beta-scaffold. E105 is predicted to interact with the LOV-STAS linker region, suggested to play a role in phototropin signaling.
Buttani
Valentina
V
Department of Physics, University of Parma, viale G.P. Usberti 7/A, Parma, Italy.
Gärtner
Wolfgang
W
Losi
Aba
A
D001412:Bacillus subtilis / Q000737:chemistry*; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D001665:Binding Sites; D006153:Guanosine Diphosphate / Q000378:metabolism*; D006157:Guanosine Monophosphate / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism*; D008027:Light; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics*; D011487:Protein Conformation; D013057:Spectrum Analysis
D001426:Bacterial Proteins; D006153:Guanosine Diphosphate; D006157:Guanosine Monophosphate; D006160:Guanosine Triphosphate