Cloning, characterization and transmission blocking potential of midgut carboxypeptidase A in Anopheles stephensi
Por:
VenkatRao V., Kumar S.K., Sridevi P., Muley V.Y., Chaitanya R.K.
Publicada:
1 ene 2017
Resumen:
Transmission-blocking vaccines (TBV) interrupt malaria parasite transmission and hence form an important component for malaria eradication. Mosquito midgut exopeptidases such as aminopeptidase N & carboxypeptidase B have demonstrated TBV potential. In the present study, we cloned and characterized carboxypeptidase A (CPA) from the midgut of an important malarial vector, Anopheles stephensi. ClustalW amino acid alignment and in silico 3-dimensional structure analysis of CPA predicted the presence of active sites involved in zinc and substrate binding that are conserved among all the known mosquito species. Real-time PCR analysis demonstrated that CPA is predominantly expressed in the midgut throughout the mosquito life cycle and that this gene is significantly elevated in P. berghei-infected mosquitoes compared to uninfected blood-fed controls. The high midgut CPA activity correlated with the prominent mRNA levels observed. Peptide-based anti-CPA antibodies were raised that cross-reacted specifically to ~48 kDa and ~37 kDa bands, which correspond to zymogen and active forms of CPA. Further, the addition of CPA-directed antibodies to P. berghei-containing blood meal significantly reduced the mosquito infection rate in the test group compared to control and blocked the parasite development in the midgut. These results support further development of A. stephensi CPA as a candidate TBV. © 2016 Elsevier B.V.
Filiaciones:
VenkatRao V.:
Department of Animal Biology, School of Life Sciences, University of Hyderabad, Hyderabad, 500046, India
Kumar S.K.:
Department of Animal Biology, School of Life Sciences, University of Hyderabad, Hyderabad, 500046, India
Sridevi P.:
Department of Biotechnology, Indira Gandhi National Tribal University, Amarkantak, 484224, India
Muley V.Y.:
Centre for Computational Science, School of Basic & Applied Sciences, Central University of Punjab, Bhatinda, 151001, India
Chaitanya R.K.:
Centre for Animal Sciences, School of Basic & Applied Sciences, Central University of Punjab, Bhatinda, 151001, India
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