ABSTRACT
Alzheimer's disease (AD) is the most common neurodegenerative disease, where ß-amyloid (Aß) plays a key role in forming conglomerated senile plaques. The receptor of advanced glycation end products (RAGE) is considered a therapeutic target since it transports Aß into the central nervous system, favoring the pathology progression. Due to the lack of effective therapies for AD, several therapeutic approaches are under development, being vaccines considered a promising alternative. Herein, the use of the Algevir system was explored to produce in the Schizochytrium sp. microalga the LTB:RAGE vaccine candidate. Algevir relies in an inducible geminiviral vector and led to yields of up to 380 µg LTB:RAGE/g fresh weight biomass at 48-h post-induction. The Schizochytrium-produced LTB:RAGE vaccine retained its antigenic activity and was highly stable up to temperatures of 60 °C. These data demonstrate the potential of Schizochytrium sp. as a platform for high production of thermostable recombinant antigens useful for vaccination against AD.
Subject(s)
Alzheimer Vaccines/metabolism , Bacterial Toxins/genetics , Enterotoxins/genetics , Escherichia coli Proteins/genetics , Microalgae/growth & development , Receptor for Advanced Glycation End Products/chemistry , Alzheimer Disease/drug therapy , Alzheimer Disease/metabolism , Alzheimer Vaccines/genetics , Alzheimer Vaccines/pharmacology , Bacterial Toxins/chemistry , Bacterial Toxins/metabolism , Cloning, Molecular , Enterotoxins/chemistry , Enterotoxins/metabolism , Epitopes/genetics , Escherichia coli/genetics , Escherichia coli/metabolism , Escherichia coli Proteins/chemistry , Escherichia coli Proteins/metabolism , Humans , Microalgae/metabolism , Protein Engineering , Receptor for Advanced Glycation End Products/antagonists & inhibitors , Receptor for Advanced Glycation End Products/genetics , Recombinant Fusion Proteins/metabolism , Recombinant Fusion Proteins/pharmacologyABSTRACT
The use of recombinant algae for the production of valuable compounds is opening promising biotechnological applications. However, the development of efficient expression approaches is still needed to expand the exploitation of microalgae in biotechnology. Herein, the concept of using viral expression vectors in microalgae was explored for the first time. An inducible geminiviral vector leading to Rep-mediated replication of the expression cassette allowed the production of antigenic proteins at high levels. This system, called Algevir, allows the production of complex viral proteins (GP1 from Zaire ebolavirus) and bacterial toxin subunits (B subunit of the heat-labile Escherichia coli enterotoxin), which retained their antigenic activity. The highest achieved yield was 1.25 mg/g fresh biomass (6 mg/L of culture), which was attained 3 days after transformation. The Algevir system allows for a fast and efficient production of recombinant proteins, overcoming the difficulties imposed by the low yields and unstable expression patterns frequently observed in stably transformed microalgae at the nuclear level; as well as the toxicity of some target proteins.