Probing the spectral signatures of orange carotenoid protein by orthogonal translation with aromatic non-canonical amino acids

Abstract

Orange Carotenoid Protein (OCP) is a water-soluble photoreceptor involved in photoprotection of cyanobacteria. The photoactive OCP contains a bound ketocarotenoid cofactor held in a protein matrix with a hydrogen bonding network. We have developed a system to replace essential residues of the photoactive OCP with non-canonical aromatic analogues that produce well-defined chemical or steric changes. Preliminary spectroscopic evaluation of the generated OCP variants demonstrates the potential of this "molecular surgery" to disentangle protein-chromophore interaction networks that are critical for photoreceptor function. In this way, the number and strength of key contacts with non-canonical amino acids could be controlled and manipulated. We have illustrated this principle here by replacing hydrogen bond donating residues with aromatic non-canonical amino acids that alter the state preference of OCP.

Keywords: Hydrogen bonding; Methanosarcina mazei pyrrolysyl-tRNA synthetase (MmPylRS); Non-canonical amino acids (ncAAs); Orange carotenoid protein (OCP); Xanthophyll photoconversion; aromatic ncAAs; site-Specific incorporation.