Dual role of the active site ‘lid’ regions of protochlorophyllide oxidoreductase in photocatalysis and plant development

DOI: 10.1111/febs.15542

Authors: Shaowei Zhang (The University of Manchester) , Alan R. F. Godwin (Wellcome Centre for Cell-Matrix Research, The University of Manchester) , Aoife Taylor (The University of Manchester) , Samantha J. O. Hardman (The University of Manchester) , Thomas A. Jowitt (Wellcome Centre for Cell‐Matrix Research, The University of Manchester) , Linus O. Johannissen (The University of Manchester) , Sam Hay (The University of Manchester) , Clair Baldock (Wellcome Centre for Cell‐Matrix Research, The University of Manchester) , Derren J. Heyes (The University of Manchester) , Nigel S. Scrutton (The University of Manchester)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: The Febs Journal , VOL 288 , PAGES 175 - 189

State: Published (Approved)
Published: August 2020
Diamond Proposal Number(s): 22724

Abstract: Protochlorophyllide oxidoreductase (POR) catalyses reduction of protochlorophyllide (Pchlide) to chlorophyllide, a light‐dependent reaction of chlorophyll biosynthesis. POR is also important in plant development as it is the main constituent of prolamellar bodies in etioplast membranes. Prolamellar bodies are highly organised, paracrystalline structures comprising aggregated oligomeric structures of POR–Pchlide–NADPH complexes. How these oligomeric structures are formed and the role of Pchlide in oligomerisation remains unclear. POR crystal structures highlight two peptide regions that form a ‘lid’ to the active site, and undergo conformational change on binding Pchlide. Here, we show that Pchlide binding triggers formation of large oligomers of POR using size exclusion chromatography. A POR ‘octamer’ has been isolated and its structure investigated by cryo‐electron microscopy at 7.7 Å resolution. This structure shows that oligomer formation is most likely driven by the interaction of amino acid residues in the highly conserved lid regions. Computational modelling indicates that Pchlide binding stabilises exposure of hydrophobic surfaces formed by the lid regions, which supports POR dimerisation and ultimately oligomer formation. Studies with variant PORs demonstrate that lid residues are involved in substrate binding and photocatalysis. These highly conserved lid regions therefore have a dual function. The lid residues position Pchlide optimally to enable photocatalysis. Following Pchlide binding, they also enable POR oligomerisation – a process that is reversed through subsequent photocatalysis in the early stages of chloroplast development.

Journal Keywords: chlorophyll biosynthesis; hydrophobic regions; oligomerisation; photocatalysis; protochlorophyllide oxidoreductase

Diamond Keywords: Photocatalysis; Photosynthesis; Enzymes

Subject Areas: Biology and Bio-materials, Chemistry

Diamond Offline Facilities: Electron Bio-Imaging Centre (eBIC)
Instruments: Krios I-Titan Krios I at Diamond

Added On: 07/01/2021 11:15

Documents:
febs.15542.pdf

Discipline Tags:

Plant science Earth Sciences & Environment Catalysis Chemistry Structural biology Life Sciences & Biotech

Technical Tags:

Microscopy Electron Microscopy (EM) Cryo Electron Microscopy (Cryo EM)