Abstract
BackgroundPhospholipase A1 is an enzyme that hydrolyzes phospholipids at the sn-1 position. It has potential applications across diverse fields including food, pharmaceutical, and biofuel industries. Although there has been increasing interest in the use of phospholipase A1 for degumming of plant oils during biodiesel production, production of recombinant phospholipase A1 has been hampered by low efficiency of gene expression and its toxicity to the host cell.ResultsWhile expression of phospholipase A1 in Escherichia coli resulted in extremely low productivity associated with inhibition of transformed cell growth, drastically higher production of functional phospholipase A1 was achieved in a cell-free protein synthesis system where enzyme expression is decoupled from cell physiology. Compared with expression in E. coli, cell-free synthesis resulted in an over 1000-fold higher titer of functional phospholipase A1. Cell-free produced phospholipase A1 was also used for successfully degumming crude plant oil.ConclusionsWe demonstrate successful production of Serratia sp. phospholipase A1 in a cell-free protein synthesis system. Including the phospholipase A1 investigated in this study, many industrial enzymes can interfere with the regular physiology of cells, making cellular production of them problematic. With the experimental results presented herewith, we believe that cell-free protein synthesis will provide a viable option for rapid production of important industrial biocatalysts.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-016-0563-5) contains supplementary material, which is available to authorized users.
Highlights
Phospholipase A1 is an enzyme that hydrolyzes phospholipids at the sn-1 position
Cell‐free synthesis of phospholipase A1 (PLA1) Based on the assumption that E. coli-based expression of Serratia PLA1 is hampered by the membrane toxicity of the expressed enzyme, we decided to investigate the use of cell-free protein synthesis as an alternative expression platform for this enzyme
Because cell-free protein synthesis reactions do not require the presence of an integral membrane, we expected that the enzymatic activity of the synthesized PLA1 would not inhibit continuous expression of this membrane-disrupting enzyme
Summary
Phospholipase A1 is an enzyme that hydrolyzes phospholipids at the sn-1 position It has potential applications across diverse fields including food, pharmaceutical, and biofuel industries. Unlike the conventional cell-based gene expression methods, cell-free protein synthesis uses protein synthesis machinery in open environments and can produce proteins by direct addition of template DNA. It offers a much faster and more flexible route to protein production [8]. Cell-free protein synthesis does not require the maintenance of cell viability and integrity of cellular membranes These unique features make cell-free protein synthesis a potentially ideal platform to produce recombinant enzymes that otherwise would interfere with the normal physiology of host cells
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
