Light-driven Na(+) pump from Gillisia limnaea: a high-affinity Na(+) binding site is formed transiently in the photocycle
| Autor: | Balashov, Sergei P, Imasheva, Eleonora S, Dioumaev, Andrei K, Wang, Jennifer M, Jung, Kwang-Hwan, Lanyi, Janos K |
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| Rok vydání: | 2014 |
| Předmět: |
Biochemistry & Molecular Biology
Molecular Sequence Data Sequence Homology Medical Biochemistry and Metabolomics Medicinal and Biomolecular Chemistry Microbial Bacterial Proteins Site-Directed Rhodopsins Amino Acid Sequence Schiff Bases Spectroscopy Aspartic Acid Binding Sites Sodium Hydrogen-Ion Concentration Photochemical Processes Recombinant Proteins Amino Acid Kinetics Amino Acid Substitution Fourier Transform Infrared Mutagenesis Biochemistry and Cell Biology Sodium-Potassium-Exchanging ATPase Flavobacteriaceae |
| Zdroj: | Biochemistry, vol 53, iss 48 |
| Popis: | A group of microbial retinal proteins most closely related to the proton pump xanthorhodopsin has a novel sequence motif and a novel function. Instead of, or in addition to, proton transport, they perform light-driven sodium ion transport, as reported for one representative of this group (KR2) from Krokinobacter. In this paper, we examine a similar protein, GLR from Gillisia limnaea, expressed in Escherichia coli, which shares some properties with KR2 but transports only Na(+). The absorption spectrum of GLR is insensitive to Na(+) at concentrations of ≤3 M. However, very low concentrations of Na(+) cause profound differences in the decay and rise time of photocycle intermediates, consistent with a switch from a "Na(+)-independent" to a "Na(+)-dependent" photocycle (or photocycle branch) at ∼60 μM Na(+). The rates of photocycle steps in the latter, but not the former, are linearly dependent on Na(+) concentration. This suggests that a high-affinity Na(+) binding site is created transiently after photoexcitation, and entry of Na(+) from the bulk to this site redirects the course of events in the remainder of the cycle. A greater concentration of Na(+) is needed for switching the reaction path at lower pH. The data suggest therefore competition between H(+) and Na(+) to determine the two alternative pathways. The idea that a Na(+) binding site can be created at the Schiff base counterion is supported by the finding that upon perturbation of this region in the D251E mutant, Na(+) binds without photoexcitation. Binding of Na(+) to the mutant shifts the chromophore maximum to the red like that of H(+), which occurs in the photocycle of the wild type. |
| Databáze: | OpenAIRE |
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