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Title: Substrate preference and oxygen requirement for cyanophycin synthesis by recombinant Escherichia coli

Author
item Solaiman, Daniel
item Ashby, Richard - Rick
item Zerkowski, Jonathan

Submitted to: Biocatalysis and Agricultural Biotechnology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/17/2011
Publication Date: 1/1/2012
Citation: Solaiman, D., Ashby, R.D., Zerkowski, J.A. 2012. Substrate preference and oxygen requirement for cyanophycin synthesis by recombinant Escherichia coli. Biocatalysis and Agricultural Biotechnology. 1(1):9-14.

Interpretive Summary: Cyanophycin (CGP) is a biopolymer consisting of amino acids. It has many potential industrial applications in the areas of water softerning, hydrogel, metal-ion chelation, and nutriceuticals. There is a need to find inexpensive ways to produce CGP. One way to improve the economics of CGP production is to invent a fermentation system that uses an inexpensive feedstock in an efficient manner without wasting it. In this study, we showed that CGP production is favoured by using glycerol – a surplus from biodiesel manufacturing. We showed also that the amounts of CGP produced can be increased by providing sufficient air in the fermentation system, and that the efficient conversion of glycerol to CGP can be improved by carefully controlling the amounts of glycerol added. The information obtained from this research lays down important foundation for further improving CGP production system, leading to cost-competitive products to benefit consumer public.

Technical Abstract: Cyanophycin (CGP) is a bacterial bioproduct having a straight-chain poly(aspartic acid) as a backbone with arginine pendant groups attached to it. It has many potential industrial applications in the areas of water softerning, hydrogel, metal-ion chelation, and nutriceuticals. Biotechnological production of CGP employs as producing strains the recombinant organisms that express heterologous cyanophycin synthase (cph) gene. A systematic study of fermentation parameters influencing CGP synthesis by a recombinant E. coli expressing a cphA of Synechocystis sp. showed that high aeration conditions as provided by 400 rpm stirrer speed and 1.0 L/min air flow in a Sixfors vessel (450 mL culture working-volume) resulted in high yields of cell biomass and crude CGP product. Glycerol substrate was found to yield 1.8-times higher crude CGP than glucose did under similar conditions. With glycerol as substrate, we found that a simplified fermentation scheme consisting of a straight 48-h fermentation at 37 deg C (without a 30-to-37 deg C temperature-shifting induction step) yielded compatible or higher amounts of crude CGP as those obtained under various temperature-shifting conditions. By studying the effects of glycerol concentration on CGP yields and analyzing glycerol consumption patterns, we demonstrated that substrate-to-product conversion could be increased by at least 15% and that costly leftover of unused substrate could be alleviated. The results provide important bases to optimize fermentative production of CGP using glycerol that may be obtained as a coproduct from biodiesel production.