Structural and Functional Studies of a Newly Grouped Haloquadratum walsbyi Bacteriorhodopsin Reveal the Acid-resistant Light-driven Proton Pumping Activity.
Retinal bound light-driven proton pumps are widespread in eukaryotic and prokaryotic organisms. Among these pumps, bacteriorhodopsin (BR) proteins cooperate with ATP synthase to convert captured solar energy into a biologically consumable form, ATP. In an acidic environment or when pumped-out protons accumulate in the extracellular region, the maximum absorbance of BR proteins shifts markedly to the longer wavelengths. These conditions affect the light-driven proton pumping functional exertion as well. In this study, wild-type crystal structure of a BR with optical stability under wide pH range from a square halophilic archaeon, Haloquadratum walsbyi (HwBR), was solved in two crystal forms. One crystal form, refined to 1.85 Å resolution, contains a trimer in the asymmetric unit, whereas another contains an antiparallel dimer was refined at 2.58 Å. HwBR could not be classified into any existing subgroup of archaeal BR proteins based on the protein sequence phylogenetic tree, and it showed unique absorption spectral stability when exposed to low pH values. All structures showed a unique hydrogen-bonding network between Arg(82) and Thr(201), linking the BC and FG loops to shield the retinal-binding pocket in the interior from the extracellular environment. This result was supported by R82E mutation that attenuated the optical stability. The negatively charged cytoplasmic side and the Arg(82)-Thr(201) hydrogen bond may play an important role in the proton translocation trend in HwBR under acidic conditions. Our findings have unveiled a strategy adopted by BR proteins to solidify their defenses against unfavorable environments and maintain their optical properties associated with proton pumping.
From the Institute of Biological Chemistry and Core Facilities for Protein Structural Analysis, Academia Sinica, Taipei 11529 and.
Min-Feng
MF
Hsu
the Department of Biochemical Science and Technology, College of Life Science, Yen Tjing Ling Industrial Research Institute, and.
Hsu-Yuan
HY
Fu
the Department of Biochemical Science and Technology, College of Life Science.
Chun-Jie
CJ
Cai
the Department of Biochemical Science and Technology, College of Life Science.
Hsiu-Pin
HP
Yi
the Department of Biochemical Science and Technology, College of Life Science, Institute of Biotechnology, College of Bio-Resources and Agriculture, National Taiwan University, Taipei 10617, Taiwan chiishen@ntu.edu.tw.
Chii-Shen
CS
Yang
From the Institute of Biological Chemistry and Core Facilities for Protein Structural Analysis, Academia Sinica, Taipei 11529 and ahjwang@gate.sinica.edu.tw.
Andrew H-J
AH
Wang
D000595:Amino Acid Sequence; D001105:Archaea / Q000378:metabolism*; D019843:Archaeal Proteins / Q000378:metabolism*; D001436:Bacteriorhodopsins / Q000378:metabolism*; D018360:Crystallography, X-Ray; D003593:Cytoplasm / Q000378:metabolism; D006860:Hydrogen Bonding; D006863:Hydrogen-Ion Concentration; D017136:Ion Transport; D008027:Light; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D055095:Optics and Photonics; D010802:Phylogeny; D015202:Protein Engineering; D011522:Protons; D017386:Sequence Homology, Amino Acid; D013056:Spectrophotometry, Ultraviolet; D055672:Static Electricity
D019843:Archaeal Proteins; D011522:Protons; D001436:Bacteriorhodopsins
bacteriorhodopsin; crystal structure; lipid cubic phase; membrane protein; proton pump; rhodopsin; site-directed mutagenesis; spectral-tuning; structure