Protonated rhodosemiquinone at the QB binding site of M265IT mutant reaction center of photosynthetic bacterium Rhodobacter sphaeroides
The 2nd electron transfer from the primary ubiquinone QA to the secondary ubiquinone QB in the reaction center (RC) from Rhodobacter sphaeroides involves protonated QB- intermediate state whose low pKa makes the direct observation impossible. Here, we replaced the native ubiquinone by low potential...
Elmentve itt :
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| Dokumentumtípus: | Cikk |
| Megjelent: |
2015
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| Sorozat: | BIOCHEMISTRY
54 No. 12 |
| doi: | 10.1021/bi501553t |
| mtmt: | 2865767 |
| Online Access: | http://publicatio.bibl.u-szeged.hu/17107 |
| Tartalmi kivonat: | The 2nd electron transfer from the primary ubiquinone QA to the secondary ubiquinone QB in the reaction center (RC) from Rhodobacter sphaeroides involves protonated QB- intermediate state whose low pKa makes the direct observation impossible. Here, we replaced the native ubiquinone by low potential rhodoquinone at the QB binding site of the M265IT mutant RC. Because the in situ midpoint redox potential of QA of this mutant was lowered about the same extent ( approximately 100 mV) as that of QB upon exchange of ubiquinone by low potential rhodoquinone, the interquinone (QA-->QB) electron transfer became energetically favorable. After subsequent saturating flash excitations, a period of two damped oscillation of the protonated rhodosemiquinone was observed. The QBH* was identified by 1) the characteristic band at 420 nm of the absorption spectrum after the 2nd flash and 2) smaller damping of the oscillation at 420 nm (due to the neutral form) than at 460 nm (attributed to the anionic form). The appearance of the neutral semiquinone was restricted to the acidic pH range indicating a functional pKa of less than 5.5, slightly higher than that of the native ubisemiquinone (pKa < 4.5) at pH 7. The analysis of the pH- and temperature dependences of the rates of the 2nd electron transfer supports the concept of pH-dependent pKa of the semiquinone at the QB binding site. The local electrostatic potential is severely modified by the strongly interacting neighboring acidic cluster and the pKa of the semiquinone is in the middle of the pH range of the complex titration. The kinetic and thermodynamic data are discussed according to the proton-activated electron transfer mechanism combined with pH-dependent functional pKa of the semiquinone at the QB site of the RC. |
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| Terjedelem/Fizikai jellemzők: | 2095-2103 |
| ISSN: | 0006-2960 |