Photosystem II (PSII) continuously attracts the attention of researchers aiming to unravel the riddle of its functioning and efficiency fundamental for all life on Earth. Besides, an increasing number of biotechnological applications have been envisaged exploiting and mimicking the unique properties of this macromolecular pigment-protein complex. The PSII organization and working principles have inspired the design of electrochemical water splitting schemes and charge separating triads in energy storage systems as well as biochips and sensors for environmental, agricultural and industrial screening of toxic compounds. An intriguing opportunity is the development of sensor devices, exploiting native or manipulated PSII complexes or ad hoc synthesized polypeptides mimicking the PSII reaction centre proteins as bio-sensing elements. This review offers a concise overview of the recent improvements in the understanding of structure and function of PSII donor side, with focus on the interactions of the plastoquinone cofactors with the surrounding environment and operational features. Furthermore, studies focused on photosynthetic proteins structure/function/dynamics and computational analyses aimed at rational design of high-quality bio-recognition elements in biosensor devices are discussed.

In the photosystem II (PSII) of oxygenic photosynthetic organisms, the reaction center (RC) core mediates the light-induced electron transfer leading to water splitting and production of reduced plastoquinone molecules. The reduction of plastoquinone to plastoquinol lowers PSII affinity for the latter and leads to its release. However, little is known about the role of protein dynamics in this process. Here, molecular dynamics simulations of the complete PSII complex embedded in a lipid bilayer have been used to investigate the plastoquinol release mechanism. A distinct dynamic behavior of PSII in the presence of plastoquinol is observed which, coupled to changes in charge distribution and electrostatic interactions, causes disruption of the interactions seen in the PSII-plastoquinone complex and leads to the "squeezing out" of plastoquinol from the binding pocket. Displacement of plastoquinol closes the second water channel, recently described in a 2.9 Å resolution PSII structure (Guskov et al. in Nat Struct Mol Biol 16:334-342, 2009), allowing to rule out the proposed "alternating" mechanism of plastoquinol-plastoquinone exchange, while giving support to the "single-channel" one. The performed simulations indicated a pivotal role of D-Ser264 in modulating the dynamics of the plastoquinone binding pocket and plastoquinol-plastoquinone exchange via its interaction with D-His252 residue. The effects of the disruption of this hydrogen bond network on the PSII redox reactions were experimentally assessed in the D site-directed mutant Ser264Lys.

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Highly stable natural scaffolds which tolerate multiple amino acid substitutions represent the ideal starting point for the application of rational redesign strategies to develop new catalysts of potential biomedical and biotechnological interest. The knottins family of disulphide-constrained peptides display the desired characteristics, being highly stable and characterized by hypervariability of the inter-cysteine loops. The potential of knottins as scaffolds for the design of novel copper-based biocatalysts has been tested by engineering a metal binding site on two different variants of an ω-conotoxin, a neurotoxic peptide belonging to the knottins family. The binding site has been designed by computational modelling and the redesigned peptides have been synthesized and characterized by optical, fluorescence, electron spin resonance and nuclear magnetic resonance spectroscopy. The novel peptides, named Cupricyclin-1 and -2, bind one Cu2+ ion per molecule with nanomolar affinity. Cupricyclins display redox activity and catalyze the dismutation of superoxide anions with an activity comparable to that of non-peptidic superoxide dismutase mimics. We thus propose knottins as a novel scaffold for the design of catalytically-active mini metalloproteins.

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