title: Development of a Recombinant Escherichia coli Strain for Overproduction of the Plant Pigment Anthocyanin.
abstract: Anthocyanins are water-soluble colored pigments found in terrestrial plants and are responsible for the red, blue, and purple coloration of many flowers and fruits. In addition to the plethora of health benefits associated with anthocyanins (cardioprotective, anti-inflammatory, antioxidant, and antiaging properties), these compounds have attracted widespread attention due to their promising potential as natural food colorants. Previously, we reported the biotransformation of anthocyanin, specifically cyanidin 3-O-glucoside (C3G), from the substrate (+)-catechin in Escherichia coli. In the present work, we set out to systematically improve C3G titers by enhancing substrate and precursor availability, balancing gene expression level, and optimizing cultivation and induction parameters. We first identified E. coli transporter proteins that are responsible for the uptake of catechin and secretion of C3G. We then improved the expression of the heterologous pathway enzymes anthocyanidin synthase (ANS) and 3-O-glycosyltransferase (3GT) using a bicistronic expression cassette. Next, we augmented the intracellular availability of the critical precursor UDP-glucose, which has been known as the rate-limiting precursor to produce glucoside compounds. Further optimization of culture and induction conditions led to a final titer of 350 mg/liter of C3G. We also developed a convenient colorimetric assay for easy screening of C3G overproducers. The work reported here constitutes a promising foundation to develop a cost-effective process for large-scale production of plant-derived anthocyanin from recombinant microorganisms.
authors.affiliation: Department of Chemical and Biological Engineering, University at Buffalo, the State University of New York, Buffalo, New York, USA.
authors.forename: Chin Giaw
authors.initials: CG
authors.lastname: Lim
authors.affiliation: Department of Chemical and Biological Engineering, University at Buffalo, the State University of New York, Buffalo, New York, USA Department of Chemical and Biological Engineering and Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York, USA.
authors.forename: Lynn
authors.initials: L
authors.lastname: Wong
authors.affiliation: Department of Chemical and Biological Engineering and Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York, USA.
authors.forename: Namita
authors.initials: N
authors.lastname: Bhan
authors.affiliation: Department of Chemical and Biological Engineering, University at Buffalo, the State University of New York, Buffalo, New York, USA.
authors.forename: Hila
authors.initials: H
authors.lastname: Dvora
authors.affiliation: Department of Chemical and Biological Engineering, University at Buffalo, the State University of New York, Buffalo, New York, USA Department of Chemical and Biological Engineering and Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York, USA.
authors.forename: Peng
authors.initials: P
authors.lastname: Xu
authors.affiliation: Department of Chemical and Biological Engineering, University at Buffalo, the State University of New York, Buffalo, New York, USA.
authors.forename: Sankaranarayanan
authors.initials: S
authors.lastname: Venkiteswaran
authors.affiliation: Department of Chemical and Biological Engineering, University at Buffalo, the State University of New York, Buffalo, New York, USA Department of Chemical and Biological Engineering and Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York, USA koffam@rpi.edu.
authors.forename: Mattheos A G
authors.initials: MA
authors.lastname: Koffas
mesh_terms: D000872:Anthocyanins / Q000096:biosynthesis*; D053898:Biosynthetic Pathways / Q000235:genetics*; D001711:Biotransformation; D002392:Catechin / Q000378:metabolism; D003124:Colorimetry / Q000379:methods; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D005285:Fermentation; D005505:Food Coloring Agents / Q000378:metabolism; D015870:Gene Expression; D060847:Metabolic Engineering; D010860:Pigments, Biological / Q000096:biosynthesis
chemical_list: D000872:Anthocyanins; D005505:Food Coloring Agents; D010860:Pigments, Biological; D002392:Catechin