کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
2020476 1069179 2013 7 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
The identification and expression of the full-length HtrA2 gene from Penaeus monodon (black tiger shrimp)
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی زیست شیمی
پیش نمایش صفحه اول مقاله
The identification and expression of the full-length HtrA2 gene from Penaeus monodon (black tiger shrimp)
چکیده انگلیسی


• We firstly report and characterise of the full-length HtrA2 gene in Penaeus monodon.
• We successfully express and purify of PmHtrA2 in E. coli strain BL21 Ril codon Plus.
• Gel exclusion chromatography indicates a high degree of uniformity in protein folding.
• Antibody specific to PmHtrA2 can detect the expression of PmHtrA2 in shrimp tissues.
• This work will further provide more focused studies on the function of PmHtrA2 in apoptosis regulation in shrimp.

HtrA2 is an apoptosis-activating protein to enhance the apoptotic process by preventing the formation of the IAP–caspase complex, thus freeing caspase to trigger the apoptosis pathway. Here, we presented the full-length sequence of HtrA2 from the black tiger shrimp (PmHtrA2). The full-length PmHtrA2 transcript was 1403 bp with a 1338 bp open reading frame encoding 445 amino acids and contains 5 conserved domains, namely, a mitochondrial targeting signal (MTS), a transmembrane (TM) domain, an IAP-binding motif (IBM), a serine protease domain, and a PDZ domain normally found in HtrA2 proteins of other organisms. The mature form of PmHtrA2 was cloned into the pET28b(+) and pET15bThio vectors, and the expression of the protein was compared in Escherichia coli BL21 DE3 and BL21 RIL (CodonPlus) strains. Greater quantities of stable and soluble PmHtrA2 were expressed as a thioredoxin fusion protein in E. coli BL21 RIL (CodonPlus) cells with the recombinant pET15bThio-PmHtrA2 vector. To investigate the expression of PmHtrA2 in shrimp, the crude proteins from several shrimp tissues were imaged by Western blot using the polyclonal antibody specific to the recombinant PmHtrA2. The expression of the 47-kDa immature PmHtrA2 protein could be detected in shrimp lysates from the gills and the muscles. This study is the first to report the full-length PmHtrA2 gene, which is functional in black tiger shrimp and will lead to more focused studies on the function of PmHtrA2 in apoptosis regulation during immune responses to viral infection in shrimp.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Protein Expression and Purification - Volume 92, Issue 2, December 2013, Pages 183–189
نویسندگان
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