Engineering photosynthetic production of L-lysine

Abstract

L-lysine and other amino acids are commonly produced through fermentation using strains of heterotrophic bacteria such as Corynebacterium glutamicum. Given the large amount of sugar this process consumes, direct photosynthetic production is intriguing alternative. In this study, we report the development of a cyanobacterium, Synechococcus sp. strain PCC 7002, capable of producing L-lysine with CO2 as the sole carbon-source. We found that heterologous expression of a lysine transporter was required to excrete lysine and avoid intracellular accumulation that correlated with poor fitness. Simultaneous expression of a feedback inhibition resistant aspartate kinase and lysine transporter were sufficient for high productivities, but this was also met with a decreased chlorophyll content and reduced growth rates. Increasing the reductant supply by using NH4+, a more reduced nitrogen source relative to NO3−, resulted in a two-fold increase in productivity directing 18% of fixed carbon to lysine. Given this advantage, we demonstrated lysine production from media formulated with a municipal wastewater treatment sidestream as a nutrient source for increased economic and environmental sustainability. Based on our results, we project that Synechococcus sp. strain PCC 7002 could produce lysine at areal productivities approaching that of sugar cane to lysine via fermentation using non-agricultural lands and low-cost feedstocks.

A New Approach on the Amino Acid Lysine Quantification by UV-Visible Spectrophotometry

“…The effect of pH on the color development in ninhydrin method was previously studied [13] for a variety of amino acid concentrations and in a restricted pH range (5.3-8.05), while in the present study was specifically analyzed for amino acid lysine and in a large pH range (2)(3)(4)(5)(6)(7)(8)(9)(10)(11). Apart from this, the ratio of Lys to Nin was also investigated and identified as the most important parameter affecting the reaction.…”

“…There is a growing demand for amino acids as value-added substances since they have many potential applications in the food, clinical, pharmaceutical and cosmetic industries, among others [1][2][3][4][5][6][7]. Amino acids have also an essential role in regulating neural activities in vivo [7] and have a great importance in all basic biological processes in the cell [8].…”

“…The motivation behind the choice of lysine (Lys), is related not only with its importance in the global market of animal feed additives but also for its necessity to children's physical growth [5,10,14,15]. The current industrial process to purify Lys solutions is based on ion-exchange multicolumn systems that require an excessive consumption of water and ammonia, with a consequent generation of environmentally harmful effluents.…”

Synthetic Biology Toolkits for Metabolic Engineering of Cyanobacteria

“…By employing the tools developed for chromosomal expression of heterologous genes, the inventory of biofuels and biochemicals produced in cyanobacteria has been greatly expanded . For instance, Atsumi et al expressed four heterologous genes in NS1 and NS2 along with an extra copy of the endogenous rbcLS in NS4 (Table ), leading to a high yield of isobutyraldehyde directly from CO 2 .…”

“…6803 ( Synechocystis 6803) . For instance, cyanobacteria have been engineered to produce fuels and fuel precursors, such as ethanol, isobutyraldehyde, butanol, alkenes, fatty acids and lipids, and the inventory of products has been expanded to value‐added chemicals for manufacturing, food, pharmaceuticals, and cosmetics, e.g., isoprene, squalene, l ‐lysine, limonene, and lactic acid . Besides the increasingly diverse products, mixotrophic growth of these photoautotrophic strains has been achieved by the installation of sugar‐utilizing pathways, thus establishing synthetic hybrid cyanobacterial platforms capable of continuous growth in diurnal conditions with significantly improved metabolic robustness .…”