Optimization of Glutathione S-Transferase as a Metabolic and Detoxification enzyme | [Abstract Book on Global Science Technology and Management Conference • 2023]

Author(s):

1. Asma Yaqoob: Department of Biochemistry, IBBB, The Islamia University of Bahawalpur, Bahawalpur, Pakistan

2. Nerum Tasneem: Department of Biochemistry, IBBB, The Islamia University of Bahawalpur, Bahawalpur, Pakistan

3. Hafiz Shahbaz Mahmood: The Faculty of Agriculture and Environment, Department of Entomology, The Islamia University of Bahawalpur, Bahawalpur, Pakistan

4. M. Asif Sajjad: The Faculty of Agriculture and Environment, Department of Entomology, The Islamia University of Bahawalpur, Bahawalpur, Pakistan

Abstract:

Glutathione S-transferase belongs to the family of multifunctional proteins and is mainly involved in the biotransformation by protecting the cell from any other harmful endogenous and exogenous compounds. It serves its role by catalyzing the conjugation reaction between the reduced glutathione with exogenous electrophilic substances. As a result, water soluble conjugate is produced that are easily metabolized and excreted. The present study was aimed to optimize glutathione S-transferase as metabolic and detoxification enzyme according to our laboratory conditions. 40mM sodium phosphate buffer (pH 7.4) was used for extraction of crude protein of Apis florea that was used for optimization. A new modified visible method for optimization of GST was developed and different parameters including ?max, concentrations of GSH and CDNB substrates, and incubation period were studied and it was found that 550nm ?max, 0.45mM GSH, 0.1mM CDNB and 0.1mM NBT concentrations and incubation for 20minutes at 37°C were optimum for visible method. UV method was also optimized and 340 nm ?max, 0.15mM GSH and 0.15mM CDNB concentration were found optimum as well as Km and Vmax values for GSH and CDNB 0.000917M, 0.000092M and 0.002194M/min, 0.000205M/min were obtained for UV method respectively. Specific activity 0.08µmoles/min/mg and 0.17µmoles/mg were calculated from ultraviolet and visible spectrophotometric enzyme assay respectively by using the crude extract. When both assaya were compared it was found that visible method (?max550) is more reliable and reproducible as compared to UV method (?max340). Up to my knowledge (literature survey) this method was first time used for quantification of glutathione S-transferase and has optimized successfully in biochemistry laboratory.

Page(s): 85-85

DOI: DOI not available

Published: Journal: Abstract Book on Global Science Technology and Management Conference, Volume: 0, Issue: 0, Year: 2023

Keywords:

GSH , Glutathione Stransferase GST , GSSG , detoxification enzymes , CDNB

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