Two-dimensional ⁶⁷Zn HYSCORE spectroscopy reveals that a Zn-bacteriochlorophyll: A _P′ dimer is the primary donor (P₈₄₀) in the type-1 reaction centers of Chloracidobacterium thermophilum

Chloracidobacterium (C.) thermophilum is a microaerophilic, chlorophototrophic species in the phylum Acidobacteria that uses homodimeric type-1 reaction centers (RC) to convert light energy into chemical energy using (bacterio)chlorophyll ((B)Chl) cofactors. Pigment analyses show that these RCs contain BChl a_P, Chl a_PD, and Zn²⁺-BChl a_P′ in the approximate ratio 7.1 : 5.4 : 1. However, the functional roles of these three different Chl species are not yet fully understood. It was recently demonstrated that Chl a_PD is the primary electron acceptor. Because Zn²⁺-(B)Chl a_P′ is present at low abundance, it was suggested that the primary electron donor might be a dimer of Zn²⁺-BChl a_P′ molecules. In this study, we utilize isotopic enrichment and high-resolution two-dimensional (2D) ¹⁴N and ⁶⁷Zn hyperfine sublevel correlation (HYSCORE) spectroscopy to demonstrate that the primary donor cation, P₈₄₀⁺, in the C. thermophilum RC is indeed a Zn²⁺-BChl a_P′ dimer. Density functional theory (DFT) calculations and the measured electron-nuclear hyperfine parameters of P₈₄₀⁺ indicate that the electron spin density on P₈₄₀⁺ is distributed nearly symmetrically over two Zn²⁺-(B)Chl a_P′ molecules as expected in a homodimeric RC. To our knowledge this is the only example of a photochemical RC in which the Chl molecules of the primary donor are metallated differently than those of the antenna.