| Cloning, expression and characterization of serine palmitoyltransferase (SPT)-like gene subunit (LCB2) from marine Emiliania huxleyi virus (Coccolithovirus) |
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| 作者姓名： | LIU Xuhong ZHENG Tianling CAI Yiqin LIU Jingwen |
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| 作者单位： | Bioengineering College of Jimei University, Xiamen 361021, China;Key Laboratory of MOE for Coast and Wetland Ecosystem, Xiamen University, Xiamen 361005, China;Bioengineering College of Jimei University, Xiamen 361021, China;Bioengineering College of Jimei University, Xiamen 361021, China |
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| 基金项目： | The National High Technology Research and Development Program of China under contract No. 2008AA09Z408, Fujian Province Nature Science Foundation, China under contract No. 2010J01261 and the Foundation for Innovative Research Team of Jimei University, China under contract No. 2010A007. |
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| 摘 要： | The authors have isolated and characterized a novel serine palmitoyltransferase(SPT)-like gene in marine Emiliania huxleyi virus(EhV-99B1).The open-reading frame(ORF) of EhV99B1-SPT encoded a protein of 496 amino acids with a calculated molecular mass of 96 kDa and Ip 6.01.The results of sequence analysis showed that there was about 31%-45% identity in amino acid sequence with other organisms.The maximum likelihood phylogenetic tree suggested that the EhV99B1-SPT gene possibly horizontally transferred from the eukaryote.Hydrophobic profiles of deduced amino acid sequences suggested a hydrophobic,globular and membrane-associated protein with five transmembrane domains(TMDs) motifs.Several potential N-linked glycosylation sites were presented in SPT.These results suggested that EhV99B1-SPT was an integral endoplasmic reticulum membrane protein.Despite lower sequence identity,the secondary and three-dimensional structures predicted showed that the "pocket" structure element composed of 2α-helices and 4βsheets was the catalytic center of this enzyme,with a typical conserved "TFTKSFG" active site in the N-terminal region and was very close to those of prokaryotic organisms.However,the N-terminal domain of EhV99B1-SPT most closely resembled the LCB2 catalysis subunit and the C-terminal domain most closely resembled the LCB1 regulatory subunit of other organisms which together formed a spherical molecule.This "chimera" was highly similar to the prokaryotic homologous SPT.For a functional identification,the EhV99B1-LCB2 subunit gene was expressed in Escherichia coli,which resulted in significant accumulation of new sphingolipid in E.coli cells.
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| 关 键 词： | 基因水平转移 催化亚基 功能鉴定 SPT 转移酶 丝氨酸 棕榈 海洋 |
| 收稿时间： | 2/3/2012 12:00:00 AM |
| 修稿时间： | 2012/5/28 0:00:00 |
Cloning, expression and characterization of serine palmitoyltransferase (SPT)-like gene subunit (LCB2) from marine Emiliania huxleyi virus (Coccolithovirus) |
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| LIU Xuhong,ZHENG Tianling,CAI Yiqin,LIU Jingwen.Cloning, expression and characterization of serine palmitoyltransferase (SPT)-like gene subunit (LCB2) from marine Emiliania huxleyi virus (Coccolithovirus)[J].Acta Oceanologica Sinica,2012,31(6):127-138. |
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| Authors: | LIU Xuhong ZHENG Tianling CAI Yiqin and LIU Jingwen |
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| Institution: | 1.Bioengineering College of Jimei University, Xiamen 361021, China2.Key Laboratory of MOE for Coast and Wetland Ecosystem, Xiamen University, Xiamen 361005, China |
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| Abstract: | The authors have isolated and characterized a novel serine palmitoyltransferase (SPT)-like gene in marine Emiliania huxleyi virus (EhV-99B1). The open-reading frame (ORF) of EhV99B1-SPT encoded a protein of 496 amino acids with a calculated molecular mass of 96 kDa and Ip 6.01. The results of sequence analysis showed that there was about 31%-45% identity in amino acid sequence with other organisms. The maximum likelihood phylogenetic tree suggested that the EhV99B1-SPT gene possibly horizontally transferred from the eukaryote. Hydrophobic profiles of deduced amino acid sequences suggested a hydrophobic, globular and membrane-associated protein with five transmembrane domains (TMDs) motifs. Several potential N-linked glycosylation sites were presented in SPT. These results suggested that EhV99B1-SPT was an integral endoplasmic reticulum membrane protein. Despite lower sequence identity, the secondary and three-dimensional structures predicted showed that the "pocket" structure element composed of 2α-helices and 4β-sheets was the catalytic center of this enzyme, with a typical conserved "TFTKSFG" active site in the N-terminal region and was very close to those of prokaryotic organisms. However, the N-terminal domain of EhV99B1-SPT most closely resembled the LCB2 catalysis subunit and the C-terminal domain most closely resembled the LCB1 regulatory subunit of other organisms which together formed a spherical molecule. This "chimera" was highly similar to the prokaryotic homologous SPT. For a functional identification, the EhV99B1-LCB2 subunit gene was expressed in Escherichia coli, which resulted in significant accumulation of new sphingolipid in E. coli cells. |
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| Keywords: | Emiliania huxleyi virus serine palmitoyltransferase (SPT) clone expressiong and characterization |
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