Rational design of photosynthetic reaction center protein maquettes

Abstract

New technologies for efficient solar-to-fuel energy conversion will help facilitate a global shift from dependence on fossil fuels to renewable energy. Nature uses photosynthetic reaction centers to convert photon energy into a cascade of electron-transfer reactions that eventually produce chemical fuel. The design of new reaction centers de novo deepens our understanding of photosynthetic charge separation and may one day allow production of biofuels with higher thermodynamic efficiency than natural photosystems. Recently, we described the multi-step electron-transfer activity of a designed reaction center maquette protein (the RC maquette), which can assemble metal ions, tyrosine, a Zn tetrapyrrole, and heme into an electron-transport chain. Here, we detail our modular strategy for rational protein design and show that the intended RC maquette design agrees with crystal structures in various states of assembly. A flexible, dynamic apo-state collapses by design into a more ordered holo-state upon cofactor binding. Crystal structures illustrate the structural transitions upon binding of different cofactors. Spectroscopic assays demonstrate that the RC maquette binds various electron donors, pigments, and electron acceptors with high affinity. We close with a critique of the present RC maquette design and use electron-tunneling theory to envision a path toward a designed RC with a substantially higher thermodynamic efficiency than natural photosystems.

Bioexcitons by Design: How Do We Get There?

“…Fortunately, critical progress has been made on both fronts in the last few years. Indeed, the recent de novo design of a variety of model proteins that bind Chl-like pigments − gives reason to hope that this side of the bioexciton design problem will in the coming years be “solved” to some workable degree of accuracy. The focus of this Perspective will instead be on the structure–spectrum problem: the challenge of accurately predicting optical spectra from structural data, allowing for a priori screening of design models for desired optical properties.…”

Design of Heme Enzymes with a Tunable Substrate Binding Pocket Adjacent to an Open Metal Coordination Site

“…Heme-binding proteins carry out a wide range of chemical reactions in biology − and directed evolution of cytochrome P450s, peroxidases, peroxygenases, globins, and other heme enzymes has led to a wealth of new catalytic activities. − The key structural feature that gives rise to this catalytic versatility is a pentacoordinate heme iron cofactor positioned adjacent to an open substrate binding pocket. De novo design efforts have taken advantage of the simplicity and designability of helical bundle scaffolds to generate porphyrin-containing catalysts. − However, this simplicity is also a limitation as such scaffolds cannot support large, open, and customizable active site pockets adjacent to the transition metal. These systems can also display considerable conformational flexibility/heterogeneity making them difficult to structurally characterize or rationally engineer, and covalent attachment of the heme cofactor may be needed to achieve selective catalysis.…”

De novo design of proteins housing excitonically coupled chlorophyll special pairs

“…numerous small molecule mimics [13][14][15][16][17] , which have provided valuable insights, but these can be labor-intensive to synthesize, overlook the role of protein matrix effects, which are important in native special pairs 5 , and lack the fine control over Chl−Chl distances and orientations needed to reproduce the precise geometries of native special pairs. De novo-designed Zn-tetrapyrrole monomer-binding proteins [18][19][20][21][22][23][24][25][26] and de novo Chl dimer proteins [27][28][29][30] have contributed to the understanding of light harvesting and quenching of excitation energy, but no Chl dimer structures have been determined experimentally in designed proteins. Systematic methods for assembling Chl dimers with predefined geometries are lacking, making it difficult to correlate structure and function, and despite decades of active research, there has been no generalizable strategy for assembling Chl dimers that precisely match special pair geometries.…”