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Biochemical Engineering Journal, Volume 68, 15 October 2012, Pages 114-119
Juan A. Moreno-Cid, Mario Canales, Jose de la Fuente
Instituto de Investigacion en Recursos Cinegeticos IREC-CSIC-UCLM-JCCM, Ronda de Toledo s/n, 13005 Ciudad Real, Spain
Department of Veterinary Pathobiology, Center for Veterinary Health Sciences, Oklahoma State University, Stillwater, OK 74078, USA
Abstract
The yeast, Pichia pastoris, has been successfully used as an efficient system to produce heterologous proteins. The secretion of the recombinant Aedes albopictus mosquito akirin (AKR) from P. pastoris using an extractive bioconversion in an aqueous two-phase system (ATPS) fed-batch fermentation process allowed the conduction of vaccination trials to assay its protective efficacy against different arthropod vector species. Herein, we report the development of a semicontinuous process for the extractive bioconversion in an ATPS coupled with a simple and easy to scale up process for polymer recycling as an effective alternative to improve production of recombinant A. albopictus AKR in P. pastoris. Six repetitive batches of the extractive bioconversion were conducted in a semicontinuous process by replacing 2/3 of the culture and recycling the top polymeric phase every 72 h. Overall process in a 5-L fermentor yielded a protein concentration in the bottom phase of 0.46 ± 0.13 mg mL−1 and 6 g of total purified protein in 490 h with a reduction of 25% of the operation time and 27% of the operation cost when compared to the fed-batch ATPS process. These results improve the production process for mosquito AKR and suggest the possibilities to develop similar processes for the expression of other recombinant proteins in P. pastoris.
Additional Information:
Towards a universal vaccine for the control of vector-borne diseases.
Vector-borne diseases are an important threat to human and animal health worldwide. Vaccination is an environmentally friendly alternative for the control of vector infestations and pathogen infection that allows control of several diseases by targeting their common vector (de la Fuente et al. 2011. Vet. Parasitol. 181, 17-22). Recombinant mosquito Akirin (AKR), tick Subolesin (SUB) and AKR/SUB chimeras containing protective epitopes from both proteins have been produced and proposed as candidate antigens for a universal vaccine against multiple arthropod vectors (de la Fuente, J. 2012. The Vet. J. 194: 139-140). Experimental vaccines containing these conserved recombinant antigens have been effective for the control of tick, mosquito and sand fly infestations (Moreno-Cid et al. 2010. Vaccine 29, 77-82; de la Fuente et al. 2011. Vet. Parasitol. 181, 17-22; Moreno-Cid et al. 2013. Vaccine 31: 1187-1196) and the infection with tick-borne pathogens (Anaplasma phagocytophilum, the causative agent of Human Granulocytic Anaplasmosis – HGA, de la Fuente et al. 2006. Parasitol. Res. 100, 85-91; Borrelia burgdorferi, the causative agent of Lyme Disease, Bensaci et al. 2012. Vaccine 30, 6040-6046; Anaplasma marginale and Babesia bigemina, the causative agents of Bovine Anaplasmosis and Bovine Babesiosis, respectively, Merino et al. 2011. Vaccine 29, 8575-8579). These results demonstrated the efficacy of SUB/AKR vaccines for the control of vectors and pathogens that affect human and animal health and suggested the possibility of vaccinating animal species that are hosts and/or affected by these vectors and pathogens. Eventually, human populations at risk could also be targeted with these vaccines. Bioprocesses that improve production yield while reducing costs are essential for the development of cost-effective vaccines for the control of vector infestations and vector-borne diseases. These vaccines need to reach both industrialized and developing countries, with special emphasis in this later group where several vector-borne diseases are endemic and cause millions of human deaths and huge losses for animal production every year.
Our group SaBio (Health and Biotechnology) at IREC focuses on research addressing health issues, with special emphasis on diseases shared between humans, domestic animals and wildlife and improving animal conservation and welfare.
José de la Fuente works at SaBio, Instituto de Investigación en Recursos Cinegéticos IREC-CSIC-UCLM-JCCM, Ciudad Real, Spain ([email protected]).
