Reconstitution of the heliobacterial photochemical reaction center and cytochrome c 553 into a proteoliposome system

William A. Johnson, Kevin E. Redding

Research output: Contribution to journalArticle

Abstract

The heliobacterial reaction center (HbRC) is the simplest known photochemical reaction center, in terms of its polypeptide composition. In the heliobacterial cells, its electron donor is a cytochrome (cyt) c553 attached to the membrane via a covalent linkage with a diacylglycerol. We have reconstituted purified HbRC into liposomes mimicking the phospholipid composition of heliobacterial membranes. We also incorporated a lipid with a headgroup containing Ni(II):nitrilotriacetate (NTA) to provide a binding site for the soluble version of the heliobacterial cyt c553 in which the N-terminal membrane attachment site is replaced by a hexahistidine tag. The HbRC was inserted into the liposomes with the donor side preferentially exposed to the exterior; this bias increased to nearly 100% with higher concentrations (≥ 10 mol%) of the Ni(II)-NTA lipid in the membrane, and is most likely due to the net negative charge of the surface of the membrane. The HbRC in proteoliposomes without the Ni(II)-NTA lipid exhibited normal charge separation and subsequent charge recombination of the P800 +FX state in 15 ms; however, the oxidized primary donor (P800 +) was not significantly reduced by added H6-cyt c553. In contrast, with proteoliposomes containing the Ni(II)-NTA lipid, addition of H6-cyt c553 resulted in a new kinetic component resulting from fast reduction (2–5 ms) of P800 + by H6-cyt c553. The contribution of this kinetic component varied with the concentration of added H6-cyt c553 and could represent 80% or more of the total P800 + decay. Thus, the HbRC and its interaction with its native electron donor have been reconstituted into an artificial membrane system.

Original languageEnglish
Pages (from-to)241-250
Number of pages10
JournalPhotosynthesis Research
Volume143
Issue number3
DOIs
Publication statusPublished - Mar 1 2020

Keywords

  • Cytochrome c
  • Heliobacteria
  • Proteoliposomes
  • Reaction center
  • Reconstitution

ASJC Scopus subject areas

  • Biochemistry
  • Plant Science
  • Cell Biology

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