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Synthesis, Conformation, and Activity of Human Insulin-Like Peptide 5 (INSL5)
Insulin-like peptide 5 (INSL5) was first identified through searches of the expressed sequence tags (EST) databases. Primary sequence analysis showed it to be a prepropeptide that was predicted to be processed in vivo to yield a two-chain sequence (A and B) that contained the insulin-like disulfide...
Autores principales: | , , , , , , , , |
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Formato: | Texto |
Lenguaje: | English |
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WILEY-VCH Verlag
2008
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2699039/ https://www.ncbi.nlm.nih.gov/pubmed/18576448 http://dx.doi.org/10.1002/cbic.200800113 |
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author | Hossain, Mohammed Akhter Bathgate, Ross A D Kong, Chze K Shabanpoor, Fazel Zhang, Suode Haugaard-Jönsson, Linda M Rosengren, K Johan Tregear, Geoffrey W Wade, John D |
author_facet | Hossain, Mohammed Akhter Bathgate, Ross A D Kong, Chze K Shabanpoor, Fazel Zhang, Suode Haugaard-Jönsson, Linda M Rosengren, K Johan Tregear, Geoffrey W Wade, John D |
author_sort | Hossain, Mohammed Akhter |
collection | PubMed |
description | Insulin-like peptide 5 (INSL5) was first identified through searches of the expressed sequence tags (EST) databases. Primary sequence analysis showed it to be a prepropeptide that was predicted to be processed in vivo to yield a two-chain sequence (A and B) that contained the insulin-like disulfide cross-links. The high affinity interaction between INSL5 and the receptor RXFP4 (GPCR142) coupled with their apparent coevolution and partially overlapping tissue expression patterns strongly suggest that INSL5 is an endogenous ligand for RXFP4. Given that the primary function of the INSL5–RXFP4 pair remains unknown, an effective means of producing sufficient quantities of this peptide and its analogues is needed to systematically investigate its structural and biological properties. A combination of solid-phase peptide synthesis methods together with regioselective disulfide bond formation were used to obtain INSL5. Both chains were unusually resistant to standard synthesis protocols and required highly optimized conditions for their acquisition. In particular, the use of a strong tertiary amidine, DBU, as N(α)-deprotection base was required for the successful assembly of the B chain; this highlights the need to consider incomplete deprotection rather than acylation as a cause of failed synthesis. Following sequential disulfide bond formation and chain combination, the resulting synthetic INSL5, which was obtained in good overall yield, was shown to possess a similar secondary structure to human relaxin-3 (H3 relaxin). The peptide was able to inhibit cAMP activity in SK-N-MC cells that expressed the human RXFP4 receptor with a similar activity to H3 relaxin. In contrast, it had no activity on the human RXFP3 receptor. Synthetic INSL5 demonstrates equivalent activity to the recombinant-derived peptide, and will be an important tool for the determination of its biological function. |
format | Text |
id | pubmed-2699039 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2008 |
publisher | WILEY-VCH Verlag |
record_format | MEDLINE/PubMed |
spelling | pubmed-26990392009-06-25 Synthesis, Conformation, and Activity of Human Insulin-Like Peptide 5 (INSL5) Hossain, Mohammed Akhter Bathgate, Ross A D Kong, Chze K Shabanpoor, Fazel Zhang, Suode Haugaard-Jönsson, Linda M Rosengren, K Johan Tregear, Geoffrey W Wade, John D Chembiochem Full Paper Insulin-like peptide 5 (INSL5) was first identified through searches of the expressed sequence tags (EST) databases. Primary sequence analysis showed it to be a prepropeptide that was predicted to be processed in vivo to yield a two-chain sequence (A and B) that contained the insulin-like disulfide cross-links. The high affinity interaction between INSL5 and the receptor RXFP4 (GPCR142) coupled with their apparent coevolution and partially overlapping tissue expression patterns strongly suggest that INSL5 is an endogenous ligand for RXFP4. Given that the primary function of the INSL5–RXFP4 pair remains unknown, an effective means of producing sufficient quantities of this peptide and its analogues is needed to systematically investigate its structural and biological properties. A combination of solid-phase peptide synthesis methods together with regioselective disulfide bond formation were used to obtain INSL5. Both chains were unusually resistant to standard synthesis protocols and required highly optimized conditions for their acquisition. In particular, the use of a strong tertiary amidine, DBU, as N(α)-deprotection base was required for the successful assembly of the B chain; this highlights the need to consider incomplete deprotection rather than acylation as a cause of failed synthesis. Following sequential disulfide bond formation and chain combination, the resulting synthetic INSL5, which was obtained in good overall yield, was shown to possess a similar secondary structure to human relaxin-3 (H3 relaxin). The peptide was able to inhibit cAMP activity in SK-N-MC cells that expressed the human RXFP4 receptor with a similar activity to H3 relaxin. In contrast, it had no activity on the human RXFP3 receptor. Synthetic INSL5 demonstrates equivalent activity to the recombinant-derived peptide, and will be an important tool for the determination of its biological function. WILEY-VCH Verlag 2008-07-21 2008-06-24 /pmc/articles/PMC2699039/ /pubmed/18576448 http://dx.doi.org/10.1002/cbic.200800113 Text en Copyright © 2008 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim http://creativecommons.org/licenses/by/2.5/ Re-use of this article is permitted in accordance with the Creative Commons Deed, Attribution 2.5, which does not permit commercial exploitation. |
spellingShingle | Full Paper Hossain, Mohammed Akhter Bathgate, Ross A D Kong, Chze K Shabanpoor, Fazel Zhang, Suode Haugaard-Jönsson, Linda M Rosengren, K Johan Tregear, Geoffrey W Wade, John D Synthesis, Conformation, and Activity of Human Insulin-Like Peptide 5 (INSL5) |
title | Synthesis, Conformation, and Activity of Human Insulin-Like Peptide 5 (INSL5) |
title_full | Synthesis, Conformation, and Activity of Human Insulin-Like Peptide 5 (INSL5) |
title_fullStr | Synthesis, Conformation, and Activity of Human Insulin-Like Peptide 5 (INSL5) |
title_full_unstemmed | Synthesis, Conformation, and Activity of Human Insulin-Like Peptide 5 (INSL5) |
title_short | Synthesis, Conformation, and Activity of Human Insulin-Like Peptide 5 (INSL5) |
title_sort | synthesis, conformation, and activity of human insulin-like peptide 5 (insl5) |
topic | Full Paper |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2699039/ https://www.ncbi.nlm.nih.gov/pubmed/18576448 http://dx.doi.org/10.1002/cbic.200800113 |
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