Molecular structure, tissue distribution and functional characterization of interferon-¦Ã-inducible lysosomal thiol reductase (GILT) gene in chicken (Gallus gallus)

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• 作者：Lin Yang^a ; Xiang Cao^a ; Xuemei Ji^a ; Hongzhen Liu^a ; Haitao Wu^a ; Wei Gu^a ; ^c ; Shuangquan Zhang^a ; ^b ; ^c ; ^{shuangquanzhang@yahoo.com} ; ^{shuangquan_z@yahoo.com}
• 关键词：cGILT ; Lipopolysaccharide (LPS) ; Real-time PCR ; Thiol reductase activity
• 刊名：Veterinary Immunology and Immunopathology
• 出版年：2013
• 出版时间：15 May, 2013
• 年：2013
• 卷：153
• 期：1-2
• 页码：140-145
• 全文大小：773 K

文摘

Interferon-¦Ã-inducible-lysosomal thiol reductase (GILT) plays a key role in the processing and presentation of MHC class II-restricted antigen (Ag) by catalyzing disulfide bond reduction, thus unfolding native protein Ag and facilitating subsequent cleavage by proteases. In this study, we reported the cloning of a GILT gene homologue from chicken (designated cGILT). The open reading frame (ORF) of cGILT consists of 762 bases, encoding a protein of 253 amino acids, with a putative molecular weight of 28 kDa. The deduced protein possesses the typical structural feature of known GILT proteins, including an active-site motif, a GILT signature sequence, and 6 conserved cysteines. Genomic analysis revealed that cGILT gene, spanning a 1868 bp fragment, contained seven exons interrupted by six introns. The result of real-time PCR showed that cGILT mRNA was expressed in a tissue-specific manner, while the cGILT mRNA expression was obviously up-regulated in spleen and PBMCs after stimulation with lipopolysaccharide (LPS). After expression as a soluble protein in Escherichia coli and purification by Ni-NTA affinity chromatography, cGILT was demonstrated to exhibit thiol reductase activity on IgG substrate.