Efficient folding of the insect neuropeptide eclosion hormone by protein disulfide isomerase.
Eclosion hormone is an insect neuropeptide that consists of 62 amino acid residues including three disulfide bonds. We have previously reported its hypothetical 3D structure consisting mainly of three alpha-helices. In this paper, we report the effects of chaperone proteins on the refolding of denatured eclosion hormone in a redox buffer containing reduced and oxidized glutathione. Urea-denatured eclosion hormone was spontaneously reactivated within 1 min with a yield of more than 90%, while beta-mercaptoethanol-denatured eclosion hormone was reactivated in a few minutes with a yield of 75%. Under the same experimental conditions, eclosion hormone treated with beta-mercaptoethanol and urea was reactivated slowly with a yield of 47% over a period of 2 h. Protein disulfide isomerase, a eucaryotic chaperone protein, markedly increased the reactivation yield and rate of the totally denatured hormone. GroE oligomers slightly improved the reactivation yield but peptidyl prolyl isomerase had no influence on yield or rate. We propose that the folding pathway of eclosion hormone involves at least two rate-limiting steps, and that protein disulfide isomerase is likely to be involved in the folding in insect neuronal cells.
Fujita
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N
Department of Biotechnology, Faculty of Science and Engineering, Ritsumeikan University, Noji-higashi, Kusatsu 525-8577, Japan. nori@se.ritsumei.ac.jp
Yoshida
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M
D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D001681:Biological Assay / Q000379:methods; D018833:Chaperonins; D029968:Escherichia coli Proteins; D006360:Heat-Shock Proteins / Q000737:chemistry / Q000378:metabolism; D007301:Insect Hormones / Q000737:chemistry* / Q000378:metabolism; D007313:Insecta; D008623:Mercaptoethanol / Q000494:pharmacology; D018832:Molecular Chaperones / Q000737:chemistry / Q000378:metabolism; D009479:Neuropeptides / Q000737:chemistry; D011489:Protein Denaturation / Q000187:drug effects; D019704:Protein Disulfide-Isomerases / Q000378:metabolism*; D017510:Protein Folding; D013329:Structure-Activity Relationship; D014508:Urea / Q000494:pharmacology
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; C028715:GroE protein, E coli; D006360:Heat-Shock Proteins; D007301:Insect Hormones; D018832:Molecular Chaperones; D009479:Neuropeptides; D008623:Mercaptoethanol; C042645:eclosion hormone; D014508:Urea; D018833:Chaperonins; D019704:Protein Disulfide-Isomerases