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Cloning, Expression and Molecular Characterization of Glutathione Transferase P1-1 Gene from the Camel, Camelus dromedarius

Cloning, Expression and Molecular Characterization of Glutathione Transferase P1-1 Gene from the Camel, Camelus dromedarius

Farid S. Ataya,1,2,* Dalia Fouad,3,4 Ajamaluddin Malik,1 Nikolaos E. Labrou,5 Mohamed S. Daoud1,6 and Hesham M. Saeed7

1Department of Biochemistry, College of Science, King Saud University, P.O. Box 2455, Riyadh 11451, Saudi Arabia 
2Molecular BiologyDepartment, Genetic Engineering Division, National Research Centre, 33 El-Bohouth St. (former El-Tahrir St.), P.O. 12622, Dokki, Giza, Egypt
3Department of Zoology, College of Science, King Saud University, P.O. 22452, Riyadh 11459, Saudi Arabia
4Department of Zoology and Entomology, Faculty of Science, Helwan University, Ein Helwan, Cairo, Egypt
5Laboratory of Enzyme Technology, Department of Biotechnology, School of Food, Biotechnology and Development, Agricultural University of Athens, 75 Iera Odos Street, GR-11855-Athens, Greece
6King Fahd Unit Laboratory, Department of Clinical and Chemical Pathology, Kasr Al-Ainy University Hospital, Cairo University, El-Manial, Cairo 11562, Egypt
7Department of Biotechnology, Institute of Graduate Studies and Research, Alexandria University, Alexandria, Egypt

*      Corresponding author: fataya@ksu.edu.sa

 

Fig. 1.

A, Agarose gel (1.2% w/v) electrophoresis for RT–PCR products of CdGSTP1-1 using the specified primers and camel liver cDNA. Left, DNA ladder; Right, RT–PCR products; B, SDS-PAGE analysis of purified CdGSTP1-1 enzyme. Lane 1, molecular weight markers; Lane 2, crude extract of E. coli BL21(DE3) uninduced; Lane 3 and 4, crude extract of E. coli BL21(DE3) induced with 1 mM IPTG after 2 and 4 h.

Fig. 2.

The nucleotide and deduced amino acid sequences of the cloned CdGSTP1-1 (GenBank accession number HM132060 and ADJ57597, respectively.

Fig. 3.

Amino acid sequence alignment of CdGSTP1-1 with homologues enzymes. The alignment was generated with the JalView program. The sequences that were used and their accession numbers are: Sus scrofa, 2GSR_A; Camelus ferus, EPY87515.1; Canis lupus familiaris, NP_001239096; Equus caballus, XP_001498156; Bos taurus, NP_803482.1; Pongo abilli, NP_001127471.1; Macaca mullata, NP_001036141.1; Mus musculus, NP_038569.1; Homo sapiens, NP_000843.1; Anolis carolinensis, XP_003215129.1; Cyprinus carpio, ABD67510.1; Danio rerio, NP_571809.1; Bufo bufo, AAN04480.1; Salmo salar, ACI70112.1; Helobdella robusta, ESO09543.1; Aplysia californica, XP_005099401.1; Mytilus galloprovincialis, AAM91994.1.

Fig. 4.

The phylogenetic tree of CdGSTP1-1 with homologues enzymes shown in Figure 3.

Fig. 5.

 Relative mRNA expression levels of CdGSTP1-1 transcript in different camel tissues using the 18S ribosomal subunit as the housekeeping gene.

Fig. 6.

Homology model of CdGSTP1-1. A, N-terminal domain (blue) and the C-terminal domain (orange); B, topology diagram for CdGSTP1-1. The N-terminal domain is colored blue and the c-terminal colored orange.

Fig. 7.

A, Superimposed modeled CdGSTP1-1 (violet) on template porcine GSTpi (yellow). The modeled CdGSTP1-1 indicated very high similarity in folding pattern with porcine GSTpi; B, surface view of the G- and H-site of CdGSTP1-1. The two subunits are shown in green and yellow. The bound inhibitor, glutathione sulfonate, is shown with magenta color; C, Comparison of the G-site binding residues and other important residues in camel (green) and porcine GSTpi (yellow).

Pakistan Journal of Zoology

October

Vol. 50, Iss. 5, Pages 1601-1998

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