Phycobilisomes Harbor FNRL in Cyanobacteria

Abstract

Cyanobacterial phycobilisomes (PBSs) are photosynthetic antenna complexes that harvest light energy and supply it to two reaction centers (RCs) where photochemistry starts. PBSs can be classified into two types, depending on the presence of allophycocyanin (APC): CpcG-PBS and CpcL-PBS. Because the accurate protein composition of CpcL-PBS remains unclear, we describe here its isolation and characterization from the cyanobacterium Synechocystis sp. strain 6803. We found that ferredoxin-NADP⁺ oxidoreductase (or FNR_L), an enzyme involved in both cyclic electron transport and the terminal step of the electron transport chain in oxygenic photosynthesis, is tightly associated with CpcL-PBS as well as with CpcG-PBS. Room temperature and low-temperature fluorescence analyses show a red-shifted emission at 669 nm in CpcL-PBS as a terminal energy emitter without APC. SDS-PAGE and quantitative mass spectrometry reveal an increased content of FNR_L and CpcC2, a rod linker protein, in CpcL-PBS compared to that of CpcG-PBS rods, indicative of an elongated CpcL-PBS rod length and its potential functional differences from CpcG-PBS. Furthermore, we combined isotope-encoded cross-linking mass spectrometry with computational protein structure predictions and structural modeling to produce an FNR_L-PBS binding model that is supported by two cross-links between K⁶⁹ of FNR_L and the N terminus of CpcB, one component in PBS, in both CpcG-PBS and CpcL-PBS (cross-link 1), and between the N termini of FNR_L and CpcB (cross-link 2). Our data provide a novel functional assembly form of phycobiliproteins and a molecular-level description of the close association of FNR_L with phycocyanin in both CpcG-PBS and CpcL-PBS.IMPORTANCE Cyanobacterial light-harvesting complex PBSs are essential for photochemistry in light reactions and for balancing energy flow to carbon fixation in the form of ATP and NADPH. We isolated a new type of PBS without an allophycocyanin core (i.e., CpcL-PBS). CpcL-PBS contains both a spectral red-shifted chromophore, enabling efficient energy transfer to chlorophyll molecules in the reaction centers, and an increased FNR_L content with various rod lengths. Identification of a close association of FNR_L with both CpcG-PBS and CpcL-PBS brings new insight to its regulatory role for fine-tuning light energy transfer and carbon fixation through both noncyclic and cyclic electron transport.

Keywords: CpcL-PBS; isotopic cross-linking; mass spectrometry; photosynthesis.