The future of ACEI
production
ACEI are a major first line treatment in managing hypertension, but
synthetic ACEI carry side-effects.
Food-derived ACEIp have been effective in BP reduction without adverse
effects and can be used as an alternative to synthetic ACEI. Potent
ACEIp have been isolated from whole-food protein hydrolysates of both
animal and plant origin. However, protein hydrolysis-based methods
generate complex peptidic mixtures from which ACEIp must be purified,
increasing production costs. Besides, the resulting recovery yields and
ACEIp bioavailability are low, rendering the process economically
unviable.
The use of recombinant technology for the production of ACEIp can extend
its application for functional foods and pharmaceutical purposes as it
allows the large-scale and low-cost production. E. coli has been
the preferred host for recombinant ACEIp production, but lactic acid
bacteria (LAB), plants (predominantly cereals and legumes) and the
microalgae C. reinhardtii have also emerged as ACEIp production
hosts.
Bacteria, such as E. coli have the advantage of growing fast and
not requiring complex culture systems. A diverse set of molecular tools
available for ACEIp production in E. coli including, a wide range
of vectors, promotors and inducers, provide also an important
competitive advantage. Furthermore, LAB species with its GRAS status and
extensive use in the dairy industry are important ACEIp production hosts
for the functional foods industry.
Nevertheless, in recent years, the potential use of plant biotechnology
for the large-scale production of pharmaceutically relevant proteins and
peptides has significantly increased. Given their low-price and safety,
plants offer many advantages for producing valuable recombinant proteins
and peptides, as ACEIp, when compared to mammalian cell cultures (Gomes
et al., 2019). Further, plants are very versatile, encompassing a wide
range of production platforms, from transgenic plants to cell suspension
cultures. The use of rice and soybean, for example, allows ACEIp
accumulation in edible seeds, a direct delivery vehicle with improved
protein stability and storage (Twyman et al., 2003), with soybeans seeds
accumulating up to 40% protein (dry weight). Legumes as soybeans
produce more proteins than other plants, being promising host systems
for molecular farming. Alternatively, tomato fruits are palatable as raw
tissue and can be lyophilized and stored for a long time (Lico et al.,
2012). The possibility of using plant tissues as direct oral delivery
means, is a major differentiating factor of bioactive peptide production
in plant platforms, compared to mainstream production platforms, such asE. coli . In bacteria, the produced peptides must undergo complex
purification procedures, and processed into a consumable product. Plant
cell suspension cultures are alternatives to transgenic plants; tobaccocalli cultures proliferate rapidly, and technologies for gene
transfer and expression are well-established for this species (Pires et
al., 2012; Schillberg et al., 2013). Further, plant cell suspension
cultures grown in sterilized contained environments provide a
cGMP-compatible production environment, advantageous to the
pharmaceutical industry and regulatory authorities (Paul et al., 2013;
Spök et al., 2008). Chloroplast-based expression platforms also attract
great interest in mass scale production of ACEIp, given its high
recombinant protein expression levels (Fletcher et al., 2007). The algae
model C. reinhardtii has fast growth rates, GRAS designation, and
can be grown in contained environments (González-Ortega et al., 2015).
Direct oral delivery of ACEIp in this host is also an option, as green
algae are edible and do not contain endotoxins, human viral or prion
contaminants (Mayfield et al., 2007). All these expression platforms are
compatible with the above described genetic engineering strategies to
improve ACEIp expression, including the multimerization of small
peptides in tandem repeats, their fusion with (or insertion in) highly
expressed proteins - with the plus of targeting/ accumulating the
ACEIp-carrying chimeric proteins in seeds - and the use of similar
chimeras, assembling different ACEI and other peptides into
multifunctional bioactive polypeptides/proteins. Together with the use
of gastrointestinal proteases cleavage sites to flank the peptides,
these approaches are perfect for the adequate release of
pharmacologically relevant peptides in edible platforms, for preventive
or therapeutic purposes.