Crystal Structure of Sensory Rhodopsin II at 2.4 Angstroms: Insights into Color Tuning and Transducer Interaction

Abstract

We report an atomic-resolution structure for a sensory member of the microbial rhodopsin family, the phototaxis receptor sensory rhodopsin II (NpSRII), which mediates blue-light avoidance by the haloarchaeon Natronobacterium pharaonis. The 2.4 angstrom structure reveals features responsible for the 70- to 80-nanometer blue shift of its absorption maximum relative to those of haloarchaeal transport rhodopsins, as well as structural differences due to its sensory, as opposed to transport, function. Multiple factors appear to account for the spectral tuning difference with respect to bacteriorhodopsin: (i) repositioning of the guanidinium group of arginine 72, a residue that interacts with the counterion to the retinylidene protonated Schiff base; (ii) rearrangement of the protein near the retinal ring; and (iii) changes in tilt and slant of the retinal polyene chain. Inspection of the surface topography reveals an exposed polar residue, tyrosine 199, not present in bacteriorhodopsin, in the middle of the membrane bilayer. We propose that this residue interacts with the adjacent helices of the cognate NpSRII transducer NpHtrII.

Steric Constraint in the Primary Photoproduct of an Archaeal Rhodopsin from Regiospecific Perturbation of C−D Stretching Vibration of the Retinyl Chromophore

“…In other words, these results strongly suggest that the counterpart of the C 14 D group is Thr204, not Tyr174. This is consistent with the X-ray structures of SRII and SRII K , where the distance from the C 14 atom changes from only 4.0 to 3.9 Å for the phenol oxygen of Tyr174, and a larger change from 4.4 Å to 3.3 Å occurs for the hydroxyl oxygen of Thr204 (7,11). It also should be noted that the peak frequency is upshifted to 2251 cm -1 for Y174F, and the broadened band implies conformational heterogeneity for the C 14 D group (spectrum e of Figure 2A).…”

“…According to the X-ray structure of SRII (11), the distance from the C 14 atom to the phenol oxygen of Tyr174 is 4.0 Å, to the hydroxyl oxygen of Thr204 is 4.4 Å, and to the hydroxyl oxygen of Thr79 is 4.3 Å (Figure 1b). The structure of SRII K shows that only the distance to Thr204 is significantly reduced, from 4.4 to 3.3 Å, by retinal photoisomerization (7,11). This result suggested that Thr204 may be the counterpart of the C 14 D group (9).…”

“…Thus, it is generally accepted that HOOP vibrations are a good probe of chromophore distortion in rhodopsins.pocket (9). In that study, all seven monodeuterated all-trans retinal analogues (positions 7,8,[10][11][12]14, and 15) were synthesized and incorporated into SRII (Figure 1a). The FTIR measurements at 77 K revealed that only the C 14 -D stretching vibration at 2244 cm -1 is significantly enhanced upon formation of the K state.…”

High-speed atomic force microscopy coming of age

“…A series of intermediates designated J, K, L, M, N, and O are defined by spectroscopy, and M (M 410 ), which has an absorption peak at 410 nm and the longest lifetime, is only the intermediate containing a deprotonated Schiff base [134]. The light-induced conformational changes in bR in the frozen activated state of the wild type (WT) and bR mutants have been studied by x-ray crystallography and electron microscopy [137][138][139]. The extent of the changes reported varies depending on the methods used, but it is generally small (0.1−0.3 nm).…”

Functional characterization of sensory rhodopsin II from Halobacterium salinarum expressed in Escherichia coli

“…Subsequently, the heterologous expression of functional pSRII from N.pharaonis has been achieved in E. coli [15] followed by expression of BR, HR and SRI from H. salinarum and purification by one step metal affinity chromatography [16,17]. Functional expression of pSRII made it possible to obtain sufficient protein for structural investigation of the receptor alone [18,19] as well as in complex with its cognate transducer [20].…”