کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
2802093 1568934 2007 9 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Molt-inhibiting hormone from the tropical land crab, Gecarcinus lateralis: Cloning, tissue expression, and expression of biologically active recombinant peptide in yeast
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی علوم غدد
پیش نمایش صفحه اول مقاله
Molt-inhibiting hormone from the tropical land crab, Gecarcinus lateralis: Cloning, tissue expression, and expression of biologically active recombinant peptide in yeast
چکیده انگلیسی

Molt-inhibiting hormone (MIH), a member of the crustacean hyperglycemic neuropeptide hormone family, inhibits ecdysteroidogenesis in the molting gland or Y-organ (YO). A cDNA encoding MIH of the land crab (Gel-MIH) was cloned from eyestalk ganglia (EG) by a combination of reverse transcriptase polymerase chain reaction (RT-PCR) and 3′- and 5′-rapid amplification of cDNA ends (RACE). The cDNA (1.4  kb) encoded MIH prohormone containing a 35 amino acid signal peptide and a 78 amino acid mature peptide. The mature peptide had the six cysteines, one glycine, two arginines, one aspartate, one phenylalanine, and one asparagine in identical positions in the highly conserved sequence characteristic of other crustacean MIHs. Gel-MIH was expressed only in the EG, as determined by RT-PCR; it was not detected in Y-organ, heart, integument, gill, testis, ovary, hepatopancreas, thoracic ganglion, or skeletal muscle. A cDNA encoding the mature peptide was used to express recombinant MIH (rMIH) using a yeast (Pichia pastoris) expression system. Two constructs were designed to yield either a mature MIH fusion protein with a c-myc epitope and histidine (His) tag at the carboxyl terminus or an untagged mature protein without the c-myc and His sequences. Immunoreactive peptides were detected in Western blots of the cell culture media with both MIH constructs, indicating secretion of the processed rMIH into the medium. Culture media containing the untagged mature peptide significantly inhibited ecdysteroid secretion by YOs from land crab and green crab (Carcinus maenas) cultured in vitro, indicating that the Gel-rMIH was biologically active.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: General and Comparative Endocrinology - Volume 150, Issue 3, February 2007, Pages 505–513
نویسندگان
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