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Molecular characterization and tissue localization of glutathione S-transferase from adult Ancylostoma ceylanicum

Published online by Cambridge University Press:  21 January 2020

J.X. Hang ,
L. He ,
A.M.I. Abuzeid ,
Y. Huang ,
Y.Q. Liu ,
X.X. Yan ,
Q. Zhao ,
X. Li ,
J.M. Liu  and
G.Q. Li Open the ORCID record for G.Q. Li [Opens in a new window]
J.X. Hang*
Affiliation:
Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou510642, China
L. He
Affiliation:
Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou510642, China
A.M.I. Abuzeid
Affiliation:
Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou510642, China
Y. Huang
Affiliation:
Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou510642, China
Y.Q. Liu
Affiliation:
Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou510642, China
X.X. Yan
Affiliation:
Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou510642, China
Q. Zhao
Affiliation:
Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou510642, China
X. Li
Affiliation:
Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou510642, China
J.M. Liu
Affiliation:
Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou510642, China
G.Q. Li
Affiliation:
Guangdong Provincial Zoonosis Prevention and Control Key Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou510642, China
*
Author for correspondence: G.Q. Li, E-mail: [email protected]
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Abstract

Glutathione S-transferases (GSTs) are a detoxifying enzyme family that is essential for parasite blood-feeding and survival, and represent potential targets for hookworm vaccine development. Multiple GST-encoding complementary DNAs (cDNAs) have been cloned from Ancylostoma caninum and Necator americanus, but there are no reports about the cloning of this enzyme from Ancylostoma ceylanicum, the animal-derived zoonotic hookworm. To study the molecular nature and tissue localization of GST of A. ceylanicum (Ace-GST), we designed primers based on the GST gene sequence of A. ceylanicum in GenBank, amplified the Ace-GST cDNA by reverse transcription polymerase chain reaction, and analysed its homology and genetic evolution relationship. The amplified product was cloned into the pET-32a vector and transformed into Escherichia coli BL21 (DE3) for expression. To prepare anti-GST polyclonal antibodies, the recombinant protein was purified and used to immunize Kunming mice. The level of immunoglobulin G (IgG) antibody in the serum of immunized mice was detected by indirect enzyme-linked immunosorbent assay, and the Ace-GST localization in adult worm was determined using the immunofluorescence method. The results showed that the full-length cDNA encoding Ace-GST was 468 bp, which had the highest homology with Ac-GST-1 (60.1%) and clustered into one branch (v-class) with Ac-GST-1 and Na-GST-1 in a phylogenetic tree. Mice immunized with recombinant Ace-GST showed specific IgG antibody response. Immunolocalization revealed that natural Ace-GST is mainly located in the epidermis, muscle and intestine of the adult. These results may lay a foundation for further studies on the biological function of Ace-GST.

Keywords

Ancylostoma ceylanicumglutathione S-transferasemolecular characterizationimmunolocalization
Type
Research Paper
Information
Journal of Helminthology , Volume 94 , 2020 , e118
Copyright
Copyright © Cambridge University Press 2020

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