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A new group of eubacterial light-driven retinal-binding proton pumps with an unusual cytoplasmic proton donor.

Biochimica et biophysica acta (2015-08-12)
Andrew Harris, Milena Ljumovic, Ana-Nicoleta Bondar, Yohei Shibata, Shota Ito, Keiichi Inoue, Hideki Kandori, Leonid S Brown
ABSTRACT

One of the main functions of microbial rhodopsins is outward-directed light-driven proton transport across the plasma membrane, which can provide sources of energy alternative to respiration and chlorophyll photosynthesis. Proton-pumping rhodopsins are found in Archaea (Halobacteria), multiple groups of Bacteria, numerous fungi, and some microscopic algae. An overwhelming majority of these proton pumps share the common transport mechanism, in which a proton from the retinal Schiff base is first transferred to the primary proton acceptor (normally an Asp) on the extracellular side of retinal. Next, reprotonation of the Schiff base from the cytoplasmic side is mediated by a carboxylic proton donor (Asp or Glu), which is located on helix C and is usually hydrogen-bonded to Thr or Ser on helix B. The only notable exception from this trend was recently found in Exiguobacterium, where the carboxylic proton donor is replaced by Lys. Here we describe a new group of efficient proteobacterial retinal-binding light-driven proton pumps which lack the carboxylic proton donor on helix C (most often replaced by Gly) but possess a unique His residue on helix B. We characterize the group spectroscopically and propose that this histidine forms a proton-donating complex compensating for the loss of the carboxylic proton donor.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
n-Dodecyl β-D-maltoside, ≥98% (GC)
Sigma-Aldrich
Nitrilotriacetic acid, Sigma Grade, ≥99%
Sigma-Aldrich
CHES, ≥99.0% (titration)
Sigma-Aldrich
Nitrilotriacetic acid, ACS reagent, ≥99.0%
Sigma-Aldrich
CHES, BioXtra, ≥99.0% (titration)
Sigma-Aldrich
n-Dodecyl β-D-maltoside, BioXtra, ≥98% (GC)
Sigma-Aldrich
Nitrilotriacetic acid, BioUltra, ≥99.0% (T)
Sigma-Aldrich
CHES, BioUltra, ≥99.5% (T)

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