Molecular cloning and expression of a putative crustacean hyperglycemic hormone of Litopenaeus vannamei in Pichia pastoris

Crustacean hyperglycemic hormone (CHH) is the most abundant and best studied member of the CHH/MIH/GIH neuropeptide hormone family. CHH plays a major role in controlling glucose levels in the hemolymph, and it also has significance in regulating molting, reproduction, and osmoregulation. In contrast...

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Autores principales:	Sánchez-Castrejón,Edna, Ponce-Rivas,Elizabeth, Aguilar,Manuel B, Díaz,Fernando
Lenguaje:	English
Publicado:	Pontificia Universidad Católica de Valparaíso 2008
Materias:	CHH Litopenaeus vannamei Pichia pastoris recombinant protein
Acceso en línea:	http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582008000400009
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spelling	oai:scielo:S0717-345820080004000092009-06-16Molecular cloning and expression of a putative crustacean hyperglycemic hormone of Litopenaeus vannamei in Pichia pastorisSánchez-Castrejón,EdnaPonce-Rivas,ElizabethAguilar,Manuel BDíaz,Fernando CHH Litopenaeus vannamei Pichia pastoris recombinant protein Crustacean hyperglycemic hormone (CHH) is the most abundant and best studied member of the CHH/MIH/GIH neuropeptide hormone family. CHH plays a major role in controlling glucose levels in the hemolymph, and it also has significance in regulating molting, reproduction, and osmoregulation. In contrast, molt-inhibiting hormone (MIH) is responsible for maintaining animals in an intermolt stage. In this study, Liv-MIH-1 cDNA, which encodes a mature neuropeptide from the eyestalk of white shrimp, Litopenaeus vannamei, was expressed in methylotrophic yeast (Pichia pastoris KM71) under the control of an alcohol oxidase promoter. Recombinant Liv-MIH-1 was secreted into the culture medium using the α-factor prepro-sequence without Glu-Ala repeats. The expected protein, which had an apparent molecular mass of 12.1 kDa, was detected by Tricine-SDS-PAGE analysis and confirmed by Western blot. Pure recombinant Liv-MIH-1 was obtained by affinity chromatography, and N-terminal sequence analysis confirmed expression of the protein. Biological assays for CHH and MIH activity were also performed. Purified recombinant Liv-MIH-1 showed the ability to elevate the glucose level of hemolymph of L. vannamei, but molting was unaffected. Since these results are in agreement with the high structural and phylogenetic similarity that has been observed between Liv-MIH-1 and other CHH neuropeptides we propose to rename the protein Liv-CHH-SG1.info:eu-repo/semantics/openAccessPontificia Universidad Católica de ValparaísoElectronic Journal of Biotechnology v.11 n.4 20082008-10-01text/htmlhttp://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582008000400009en10.4067/S0717-34582008000400009
institution	Scielo Chile
collection	Scielo Chile
language	English
topic	CHH Litopenaeus vannamei Pichia pastoris recombinant protein
spellingShingle	CHH Litopenaeus vannamei Pichia pastoris recombinant protein Sánchez-Castrejón,Edna Ponce-Rivas,Elizabeth Aguilar,Manuel B Díaz,Fernando Molecular cloning and expression of a putative crustacean hyperglycemic hormone of Litopenaeus vannamei in Pichia pastoris
description	Crustacean hyperglycemic hormone (CHH) is the most abundant and best studied member of the CHH/MIH/GIH neuropeptide hormone family. CHH plays a major role in controlling glucose levels in the hemolymph, and it also has significance in regulating molting, reproduction, and osmoregulation. In contrast, molt-inhibiting hormone (MIH) is responsible for maintaining animals in an intermolt stage. In this study, Liv-MIH-1 cDNA, which encodes a mature neuropeptide from the eyestalk of white shrimp, Litopenaeus vannamei, was expressed in methylotrophic yeast (Pichia pastoris KM71) under the control of an alcohol oxidase promoter. Recombinant Liv-MIH-1 was secreted into the culture medium using the α-factor prepro-sequence without Glu-Ala repeats. The expected protein, which had an apparent molecular mass of 12.1 kDa, was detected by Tricine-SDS-PAGE analysis and confirmed by Western blot. Pure recombinant Liv-MIH-1 was obtained by affinity chromatography, and N-terminal sequence analysis confirmed expression of the protein. Biological assays for CHH and MIH activity were also performed. Purified recombinant Liv-MIH-1 showed the ability to elevate the glucose level of hemolymph of L. vannamei, but molting was unaffected. Since these results are in agreement with the high structural and phylogenetic similarity that has been observed between Liv-MIH-1 and other CHH neuropeptides we propose to rename the protein Liv-CHH-SG1.
author	Sánchez-Castrejón,Edna Ponce-Rivas,Elizabeth Aguilar,Manuel B Díaz,Fernando
author_facet	Sánchez-Castrejón,Edna Ponce-Rivas,Elizabeth Aguilar,Manuel B Díaz,Fernando
author_sort	Sánchez-Castrejón,Edna
title	Molecular cloning and expression of a putative crustacean hyperglycemic hormone of Litopenaeus vannamei in Pichia pastoris
title_short	Molecular cloning and expression of a putative crustacean hyperglycemic hormone of Litopenaeus vannamei in Pichia pastoris
title_full	Molecular cloning and expression of a putative crustacean hyperglycemic hormone of Litopenaeus vannamei in Pichia pastoris
title_fullStr	Molecular cloning and expression of a putative crustacean hyperglycemic hormone of Litopenaeus vannamei in Pichia pastoris
title_full_unstemmed	Molecular cloning and expression of a putative crustacean hyperglycemic hormone of Litopenaeus vannamei in Pichia pastoris
title_sort	molecular cloning and expression of a putative crustacean hyperglycemic hormone of litopenaeus vannamei in pichia pastoris
publisher	Pontificia Universidad Católica de Valparaíso
publishDate	2008
url	http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582008000400009
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Molecular cloning and expression of a putative crustacean hyperglycemic hormone of Litopenaeus vannamei in Pichia pastoris

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