Optimizing multi-step B-side charge separation in photosynthetic reaction centers from Rhodobacter capsulatus.
Biochim Biophys Acta
; 1857(2): 150-159, 2016 Feb.
Article
em En
| MEDLINE
| ID: mdl-26658355
ABSTRACT
Using high-throughput methods for mutagenesis, protein isolation and charge-separation functionality, we have assayed 40 Rhodobacter capsulatus reaction center (RC) mutants for their P(+)QB(-) yield (P is a dimer of bacteriochlorophylls and Q is a ubiquinone) as produced using the normally inactive B-side cofactors BB and HB (where B is a bacteriochlorophyll and H is a bacteriopheophytin). Two sets of mutants explore all possible residues at M131 (M polypeptide, native residue Val near HB) in tandem with either a fixed His or a fixed Asn at L181 (L polypeptide, native residue Phe near BB). A third set of mutants explores all possible residues at L181 with a fixed Glu at M131 that can form a hydrogen bond to HB. For each set of mutants, the results of a rapid millisecond screening assay that probes the yield of P(+)QB(-) are compared among that set and to the other mutants reported here or previously. For a subset of eight mutants, the rate constants and yields of the individual B-side electron transfer processes are determined via transient absorption measurements spanning 100 fs to 50 µs. The resulting ranking of mutants for their yield of P(+)QB(-) from ultrafast experiments is in good agreement with that obtained from the millisecond screening assay, further validating the efficient, high-throughput screen for B-side transmembrane charge separation. Results from mutants that individually show progress toward optimization of P(+)HB(-)âP(+)QB(-) electron transfer or initial P*âP(+)HB(-) conversion highlight unmet challenges of optimizing both processes simultaneously.
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01-internacional
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MEDLINE
Assunto principal:
Feofitinas
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Fotossíntese
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Bacterioclorofilas
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Rhodobacter capsulatus
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Ubiquinona
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Complexos de Proteínas Captadores de Luz
Idioma:
En
Revista:
Biochim Biophys Acta
Ano de publicação:
2016
Tipo de documento:
Article