Protonation State of Glu142 Differs in the Green- and Blue-Absorbing Variants of Proteorhodopsin super([dagger])

• Published in: Biochemistry (Easton) 2008-01, Vol.47 (11), p.3447-3453
• Main Authors: Bergo, Vladislav B, Spudich, John L, Spudich, Elena N, Kralj, Joel M, Amsden, Jason J, Rothschild, Kenneth J
• Format: Article
• Language: English
• ISSN: 0006-2960
• DOI: 10.1021/bi7018964

Proteorhodopsins are a recently discovered class of microbial rhodopsins, ubiquitous in marine bacteria. Over 4000 variants have thus far been discovered, distributed throughout the oceans of the world. Most variants fall into one of two major groups, green- or blue-absorbing proteorhodopsin (GPR and BPR, respectively), on the basis of both the visible absorption maxima (530 versus 490 nm) and photocycle kinetics ( similar to 20 versus similar to 200 ms). For a well-studied pair, these differences appear to be largely determined by the identity of a single residue at position 105 (leucine/GPR and glutamine/BPR). We find using a combination of visible and infrared spectroscopy that a second difference is the protonation state of a glutamic acid residue located at position 142 on the extracellular side of helix D. In BPR, Glu142 (the GPR numbering system is used) is deprotonated and can act as an alternate proton acceptor, thus explaining the earlier observations that neutralization of the Schiff base counterion, Asp97, does not block the formation of the M intermediate. In contrast, Glu142 in GPR is protonated and cannot act in this state as an alternate proton acceptor for the Schiff base. On the basis of these findings, a mechanism is proposed for proton pumping in BPR. Because the pK sub(a) of Glu142 is near the pH of its native marine environment, changes in pH may act to modulate its function in the cell.