Study on the constructions and activities of three novel hGHRH analogs with N-terminal prolyl modulation+.
Growth hormone releasing hormone is one of the hormones secreted by the hypothalamus. Because of its potential applications in agriculture and medicine, its short half-life and its expensive chemical synthesis, an analog with high GHRH activity and prolonged half-life has been looked for. The fusion partner gene with 127 amino acid residues of the C-terminus from L-asparaginase was recombined respectively with asp-pro-pro-hGHRH(1-44), asp-pro-hGHRH(1-44) or asp-1pro-GHRH(2-44) genes synthesized by PCR method to form three kinds of fusion proteins with unique acid labile linker Asp-Pro. The Pro-Pro-hGHRH(1-44), Pro-hGHRH(1-44), and 1Pro-GHRH(2-44) peptides were purified to homogeneity by means of cell disruption, washing of inclusion body, ethanol fraction precipitation, acid hydrolysis, SP-Sephadex C-25 and Sephadex G-10 column chromatography. The peptide molecular mass of 5235, 5139 or 4975 Da was determined by ESI mass spectroscopy and purity was determined by SDS-PAGE. In the study of in vitro activity, the antiserum kit against human GH and peptide doses of 0.1, 1.0 and 10 microg/ml were used. These peptides obviously increased GH releases both from human pituitary and from rat pituitary. The activity comparisons showed that there was significant difference between Pro-Pro-hGHRH(1-44)-Gly-Gly-Cys and Pro-Pro-hGHRH(1-44) at 1.0 microg/ml, or between 1Pro-hGHRH(2-44) and Pro-Pro-hGHRH(1-44) or Pro-hGHRH(1-44) at 10 microg/ml. The structure-activity relationships showed that at the original C-terminus, for rat pituitary the activity of the GHRH analog with 1Tyr-->Pro was more than that of Pro-Pro-hGHRH(1-44) or Pro-hGHRH(1-44). The results showed that the analogs had good GH-releasing activity and species specificity.
Tang
Song-Shan
SS
Department of Biopharmaceutics, Guangdong Pharmaceutical University, Guangdong [corrected] China.
Du
Ming-Hua
MH
Zhang
Xiao-Wen
XW
Zhang
Juan-Hui
JH
Kong
Xin
X
Wu
Jie
J
Liu
Jing-Jing
JJ
D013006:Growth Hormone / Q000378:metabolism; D013007:Growth Hormone-Releasing Hormone / Q000031:analogs & derivatives* / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D006728:Hormones / Q000378:metabolism; D006801:Humans; D008970:Molecular Weight; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D010902:Pituitary Gland / Q000378:metabolism; D011392:Proline / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology; D013329:Structure-Activity Relationship
D006728:Hormones; D010446:Peptide Fragments; C511233:Pro-GHRH(2-44) peptide, human; C511232:Pro-Pro-hGHRH(1-44) peptide, human; C497663:Pro-Pro-hGHRH(1-44)-Gly-Gly-Cys; D011993:Recombinant Fusion Proteins; D013006:Growth Hormone; D013007:Growth Hormone-Releasing Hormone; D011392:Proline