Abstract
Adenosine triphosphate (ATP), as a universal energy currency, takes a central role in many biochemical reactions with potential for the synthesis of numerous high-value products. However, the high cost of ATP limits industrial ATP-dependent enzyme-catalyzed reactions. Here, we investigated the effect of cell-surface display of phosphotransferase on ATP regeneration in recombinant Escherichia coli. By N-terminal fusion of the super-folder green fluorescent protein (sfGFP), we successfully displayed the phosphotransferase of Pseudomonas brassicacearum (PAP-Pb) on the surface of E. coli cells. The catalytic activity of sfGFP-PAP-Pb intact cells was 2.12 and 1.47 times higher than that of PAP-Pb intact cells, when the substrate was AMP and ADP, respectively. The conversion of ATP from AMP or ADP were up to 97.5% and 80.1% respectively when catalyzed by the surface-displayed enzyme at 37 °C for only 20 min. The whole-cell catalyst was very stable, and the enzyme activity of the whole cell was maintained above 40% after 40 rounds of recovery. Under this condition, 49.01 mg/mL (96.66 mM) ATP was accumulated for multi-rounds reaction. This ATP regeneration system has the characteristics of low cost, long lifetime, flexible compatibility, and great robustness.
Highlights
Adenosine-5’-triphosphate (ATP), is one of the essential molecules in living systems [1], plays a central role in many biochemical reactions, and has the potential to synthesize many high-value products [2,3]
The synthesis and consumption of ATP play an important role in many aspects of cell metabolism, such as active transport mechanisms [1,2], ATP-binding cassette (ABC) transporters [4], and ATP as the precursor to synthesize DNA, RNA, and NAD(P) [5]
ATP is necessary for the biosynthesis of cyclic adenosine monophosphate, that is a significant second messenger in signal transduction; ATP can serve as a signal ligand for ATP-sensitive or purinergic ionotropic and G-protein coupled receptors [6]
Summary
Adenosine-5’-triphosphate (ATP), is one of the essential molecules in living systems [1], plays a central role in many biochemical reactions, and has the potential to synthesize many high-value products [2,3]. ATP provides energy for the biosynthesis of a large number of biological compounds, such as amino acids, proteins, and lipids [6]. Traditional ATP synthesis is chemically synthesized from the substrate AMP or ADP with compounds with phosphoric acid groups as the phosphorylation reagents [10]. ATP was regenerated by the direct transfer of a phosphoryl group of another phosphorylated compound to ADP and AMP for energy metabolite [2]; this is the most significant way of ATP regeneration for anaerobic microorganisms and cells during anoxia [2]
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