FT-IR difference spectroscopy elucidates crucial interactions of sensory rhodopsin I with the cognate transducer HtrI.
The phototaxis receptor sensory rhodopsin I (SRI) from Halobacterium salinarum interacts with its cognate transducer (HtrI) forming a transmembrane complex. After light excitation of the chromophore all-trans retinal, SRI undergoes structural changes that are ultimately transmitted to HtrI. The interaction of SRI with HtrI results in the closure of the receptor's proton pathway, which renders the photocycle recovery kinetics of SRI pH-independent. We demonstrate on heterologously expressed and reconstituted SRI-HtrI fusion proteins that the transmembrane part of HtrI (residues 1-52) as well as the downstream cytoplasmic part (residues 53-147) exhibit conformational changes after light excitation. The sum of these conformational changes is similar to those observed in the fusion constructs SRI-HtrI 1-71 and SRI-HtrI 1-147, which display pH-independent receptor kinetics. These results indicate the occurrence of spatially distinct conformational changes that are required for functional signal transmission. Kinetic and spectroscopic analysis of HtrI point mutants of Asn53 provides evidence that this residue is involved in the receptor-transducer interaction. We suggest that Asn53 plays a role similar to that of Asn74 of the HtrII from Natronobacterium pharaonis, the latter forming a hydrogen bond to the receptor within the membrane.
Mironova
Olga S
OS
Research Center Jülich, Institute for Structural Biology (IBI-2), 52425 Jülich, Germany.
Budyak
Ivan L
IL
Büldt
Georg
G
Schlesinger
Ramona
R
Heberle
Joachim
J
D019843:Archaeal Proteins / Q000737:chemistry* / Q000235:genetics / Q000528:radiation effects; D001216:Asparagine / Q000737:chemistry / Q000235:genetics; D025602:Halorhodopsins / Q000737:chemistry* / Q000235:genetics / Q000528:radiation effects; D008027:Light; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000528:radiation effects; D017354:Point Mutation; D025941:Protein Interaction Mapping; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000528:radiation effects; D025603:Sensory Rhodopsins / Q000737:chemistry* / Q000235:genetics / Q000528:radiation effects; D017550:Spectroscopy, Fourier Transform Infrared
D019843:Archaeal Proteins; D025602:Halorhodopsins; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; C061497:SRI protein, Halobacterium; D025603:Sensory Rhodopsins; C082504:htrI protein, Halobacterium salinarium; D001216:Asparagine