The growing demand for food and biofuels urges the vegetable oil processing industry to adopt cleaner technologies to mitigate the environmental pollution caused by chemical refining processes. Over the past decade, several enzymatic methods have proven to be efficient at reducing the generated waste, but improving the benefit-cost ratio is still necessary for the widespread adoption of this technology. In this work, we show that lecithin:cholesterol acyltransferase from Aeromonas enteropelogenes (LCAT(AE)) provides a higher extra-yield of soybean oil than a type A1 phospholipase (PLA) enzyme currently commercialized for soybean oil deep degumming. Our model indicates that crude soybean oil treated with the new enzyme generates 87% more neutral oil from phospholipids than the widely used PLA, with the corresponding reduction in waste and byproducts generation. The refined oil retains the phytosterols naturally present in crude oil, enriching its nutritional value. The results presented here position LCAT(AE) as a promising candidate to provide the green solutions needed by the industrial oil processing sector. Key points Selected LCAT gene candidates were expressed in E. coli. Aeromonas enteropelogenes LCAT hydrolyzes all the phospholipids present in crude soybean oil. The LCAT enzyme provides a higher yield of neutral oil than commercial PLA enzymes and generates less waste. The degummed oil retains sterols with high nutritional value.
        
Title: Total synthesis of long DNA sequences: synthesis of a contiguous 32-kb polyketide synthase gene cluster Kodumal SJ, Patel KG, Reid R, Menzella HG, Welch M, Santi DV Ref: Proc Natl Acad Sci U S A, 101:15573, 2004 : PubMed
To exploit the huge potential of whole-genome sequence information, the ability to efficiently synthesize long, accurate DNA sequences is becoming increasingly important. An approach proposed toward this end involves the synthesis of approximately 5-kb segments of DNA, followed by their assembly into longer sequences by conventional cloning methods [Smith, H. O., Hutchinson, C. A., III, Pfannkoch, C. & Venter, J. C. (2003) Proc. Natl. Acad. Sci. USA 100, 15440-15445]. The major current impediment to the success of this tactic is the difficulty of building the approximately 5-kb components accurately, efficiently, and rapidly from short synthetic oligonucleotide building blocks. We have developed and implemented a strategy for the high-throughput synthesis of long, accurate DNA sequences. Unpurified 40-base synthetic oligonucleotides are built into 500- to 800-bp "synthons" with low error frequency by automated PCR-based gene synthesis. By parallel processing, these synthons are efficiently joined into multisynthon approximately 5-kb segments by using only three endonucleases and "ligation by selection." These large segments can be subsequently assembled into very long sequences by conventional cloning. We validated the approach by building a synthetic 31,656-bp polyketide synthase gene cluster whose functionality was demonstrated by its ability to produce the megaenzyme and its polyketide product in Escherichia coli.