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Affinity purification, identification, and biochemical characterization of Gamma-glutamyl transpeptidase, a membrane anchored enzyme of Gigantocotyle explanatum

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Abstract

Gamma-glutamyl transpeptidase is an enzyme that facilitates the transfer of glutamyl groups from glutamyl peptides to other peptides or water. Additionally, it also participates in important processes such as amino acid transport, cellular redox control, drug detoxification, apoptosis, and DNA fragmentation in a various organism. In the present study, GGT activity in Gigantocotyle explanatum was examined in order to characterize the enzyme in the helminth system. GGT is isolated using membrane solubilization and purified through affinity column chromatography (Con-A Sepharose column). Km and Vmax values, as well as the optimal pH, optimal temperature, and incubation period, are also determined using enzyme kinetics. The hetero-dimeric property of the enzyme is demonstrated by the purified GGT, which yielded two subunits of 65.5 and 55 kDa. The optimal pH and temperature are found to be 8.0 and 37 °C, respectively. While assessing the optimal incubation time of the enzyme, it was observed that the purified GGT not only retained its functional integrity up to 15 min but also reflected considerable thermostability at higher temperatures, by retaining 78% and 25% of its initial activities at 50 °C and 60 °C, respectively. One millimolar concentration of 6-Diazo-5-Oxo Nor-isoleucine (DON), a specific inhibitor of GGT, completely abolished GGT activity. These results suggest that GGT in these worms is a catalytically active enzyme with distinguishing characteristics that can be used for further study to comprehend its function in amphistome biology and in host-parasite relationships, especially since the potential therapeutic candidacy of the GGT enzyme has already been indicated in these groups of organisms.

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Funding

All the authors gratefully acknowledge the financial support provided by the University Grant Commission, Govt. of India, New Delhi, India.

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Authors and Affiliations

  1. Section of Parasitology, Department of Zoology, Aligarh Muslim University, Uttar Pradesh, Aligarh, 202002, India

    Faiza Farhat & S. M. A. Abidi

  2. College of Medicine, Gachon University, Incheon, South Korea

    Sobia Wasim

  3. Department of Lymphoma/Myeloma, MD Anderson Cancer Center, The University of Texas, Houston, TX, USA

    Lubna Rehman

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  1. Faiza Farhat
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  2. Sobia Wasim
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  3. Lubna Rehman
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  4. S. M. A. Abidi
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Contributions

Faiza Farhat: experimental work, data curation, and writing—original draft preparation; Sobia Wasim: data analysis, reviewing, and editing; Lubna Rehman: formal analyses, proofreading, and editing; S. M. A. Abidi: conceptualization and supervision.

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Correspondence to Faiza Farhat.

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Farhat, F., Wasim, S., Rehman, L. et al. Affinity purification, identification, and biochemical characterization of Gamma-glutamyl transpeptidase, a membrane anchored enzyme of Gigantocotyle explanatum. Parasitol Res 122, 915–926 (2023). https://doi.org/10.1007/s00436-023-07786-7

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