Journal article

Translational incorporation of L-3,4-dihydroxyphenylalanine into proteins

K Ozawa, MJ Headlam, D Mouradov, SJ Watt, JL Beck, KJ Rodgers, RT Dean, T Huber, G Otting, NE Dixon

FEBS JOURNAL | WILEY | Published : 2005

Abstract

An Escherichia coli cell-free transcription/translation system was used to explore the high-level incorporation of L-3,4-dihydroxyphenylalanine (DOPA) into proteins by replacing tyrosine with DOPA in the reaction mixtures. ESI-MS showed specific incorporation of DOPA in place of tyrosine. More than 90% DOPA incorporation at each tyrosine site was achieved, allowing the recording of clean 15N-HSQC NMR spectra. A redox-staining method specific for DOPA was shown to provide a sensitive and generally applicable method for assessing the cell-free production of proteins. Of four proteins produced in soluble form in the presence of tyrosine, two resulted in insoluble aggregates in the presence of h..

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University of Melbourne Researchers