DOI: 10.1101/456087Oct 29, 2018Paper

Cell-free soluble expression of the membrane protein PsbS

BioRxiv : the Preprint Server for Biology
Maithili KrishnanAnjali Pandit

Abstract

Photosystem II subunit S (PsbS) is a membrane protein that plays an exclusive role in non-photochemical quenching for photoprotection of plants under high-light conditions. The activation mechanism of PsbS and its pH-induced conformational changes are currently unknown. For structural investigation of PsbS, effective synthesis of PsbS with selective isotope or electron-spin labels or non-natural amino acids incorporated would be a great asset. This communication presents cell-free expression as a successful method for in vitro production of PsbS that would allow such incorporation. We have optimized the cell-free method to yield soluble PsbS of ~500 ng/microl using a continuous-exchange method at 30 0C, along with a successful purification and refolding of PsbS in n-Dodecyl b-D-maltoside (b-DM) detergent. We expect that the presented protocols are transferrable for in vitro expression of other membrane proteins of the Light-Harvesting Complex family.

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