Purification and functional characterization of a protein: Bombyx mori human growth hormone like protein in silkworm pupa.
Human growth hormone (hGH) is a peptide hormone secreted by eosinophils of the human anterior pituitary, and a regulatory factor for a variety of metabolic pathways. A 30-kD protein from the pupa stage of silkworm was detected by Western blotting and confirmed by immunoprecipitation based on its abi...
| Main Authors: | , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Public Library of Science (PLoS)
2014-01-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC4254979?pdf=render |
| _version_ | 1818174010922369024 |
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| author | Jianqing Chen Tejun Shu Zhengbing Lv Zuoming Nie Jian Chen Hao Chen Wei Yu Qijing Gai Yaozhou Zhang |
| author_facet | Jianqing Chen Tejun Shu Zhengbing Lv Zuoming Nie Jian Chen Hao Chen Wei Yu Qijing Gai Yaozhou Zhang |
| author_sort | Jianqing Chen |
| collection | DOAJ |
| description | Human growth hormone (hGH) is a peptide hormone secreted by eosinophils of the human anterior pituitary, and a regulatory factor for a variety of metabolic pathways. A 30-kD protein from the pupa stage of silkworm was detected by Western blotting and confirmed by immunoprecipitation based on its ability to bind to anti-hGH antibody. This protein, named BmhGH-like protein, was purified from fresh silkworm pupas through low-temperature homogenization, filtration, and centrifugation to remove large impurity particles. The supernatants were precipitated, resuspended, and passed through a molecular sieve. Further purification by affinity chromatography and two-dimensional electrophoresis resulted in pure protein for analysis by MS MALDI-TOF-MS analysis. An alignment with predicted proteins indicated that BmhGH-like protein consisted of two lipoproteins, which we named hGH-L1 and hGH-L2. These proteins belong to the β-trefoil superfamily, with β domains similar to the spatial structure of hGH. Assays with K562 cells demonstrated that these proteins could promote cell division in vitro. To further validate the growth-promoting effects, hGH-L2 was cloned from pupa cDNA to create recombinant silkworm baculovirus vBmNPV-hGH-L2, which was used to infect silkworm BmN cells at low titer. Flow cytometric analysis demonstrated that the protein shortened the G0/G1 phase of the cells, and enabled the cells to rapidly traverse the G1/S phase transition point to enter S phase and promote cell division. Discovery of hGH-like protein in silkworm will once again arouse people's interest in the potential medicinal value of silkworm and establish the basis for the development of new hormone drugs. |
| first_indexed | 2024-12-11T19:37:36Z |
| format | Article |
| id | doaj.art-ac84064066bd4edfb3f79e3a3c0b16ba |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-11T19:37:36Z |
| publishDate | 2014-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-ac84064066bd4edfb3f79e3a3c0b16ba2022-12-22T00:53:07ZengPublic Library of Science (PLoS)PLoS ONE1932-62032014-01-01912e11435110.1371/journal.pone.0114351Purification and functional characterization of a protein: Bombyx mori human growth hormone like protein in silkworm pupa.Jianqing ChenTejun ShuZhengbing LvZuoming NieJian ChenHao ChenWei YuQijing GaiYaozhou ZhangHuman growth hormone (hGH) is a peptide hormone secreted by eosinophils of the human anterior pituitary, and a regulatory factor for a variety of metabolic pathways. A 30-kD protein from the pupa stage of silkworm was detected by Western blotting and confirmed by immunoprecipitation based on its ability to bind to anti-hGH antibody. This protein, named BmhGH-like protein, was purified from fresh silkworm pupas through low-temperature homogenization, filtration, and centrifugation to remove large impurity particles. The supernatants were precipitated, resuspended, and passed through a molecular sieve. Further purification by affinity chromatography and two-dimensional electrophoresis resulted in pure protein for analysis by MS MALDI-TOF-MS analysis. An alignment with predicted proteins indicated that BmhGH-like protein consisted of two lipoproteins, which we named hGH-L1 and hGH-L2. These proteins belong to the β-trefoil superfamily, with β domains similar to the spatial structure of hGH. Assays with K562 cells demonstrated that these proteins could promote cell division in vitro. To further validate the growth-promoting effects, hGH-L2 was cloned from pupa cDNA to create recombinant silkworm baculovirus vBmNPV-hGH-L2, which was used to infect silkworm BmN cells at low titer. Flow cytometric analysis demonstrated that the protein shortened the G0/G1 phase of the cells, and enabled the cells to rapidly traverse the G1/S phase transition point to enter S phase and promote cell division. Discovery of hGH-like protein in silkworm will once again arouse people's interest in the potential medicinal value of silkworm and establish the basis for the development of new hormone drugs.http://europepmc.org/articles/PMC4254979?pdf=render |
| spellingShingle | Jianqing Chen Tejun Shu Zhengbing Lv Zuoming Nie Jian Chen Hao Chen Wei Yu Qijing Gai Yaozhou Zhang Purification and functional characterization of a protein: Bombyx mori human growth hormone like protein in silkworm pupa. PLoS ONE |
| title | Purification and functional characterization of a protein: Bombyx mori human growth hormone like protein in silkworm pupa. |
| title_full | Purification and functional characterization of a protein: Bombyx mori human growth hormone like protein in silkworm pupa. |
| title_fullStr | Purification and functional characterization of a protein: Bombyx mori human growth hormone like protein in silkworm pupa. |
| title_full_unstemmed | Purification and functional characterization of a protein: Bombyx mori human growth hormone like protein in silkworm pupa. |
| title_short | Purification and functional characterization of a protein: Bombyx mori human growth hormone like protein in silkworm pupa. |
| title_sort | purification and functional characterization of a protein bombyx mori human growth hormone like protein in silkworm pupa |
| url | http://europepmc.org/articles/PMC4254979?pdf=render |
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