Abstract
A novel fluorescent protein termed hmGFP homologous to the green fluorescent protein (GFP) from Aequorea victoria was cloned from the tentacles of sea anemone Heteractis magnifica by EST sequencing and analysis of cDNA library and followed by using RT-PCR. The sequence analysis suggested that the chromophore, consensus amino acids, and secondary structure of 11 β-strands of hmGFP were similar to those of GFP from other species. The recombinant hmGFP protein with high purity was obtained by the fusion expression of pETTRX-hmGFP in Escherichia coli and subsequent purification. The pH sensitivity and fluorescence spectroscopy of recombinant hmGFP were characterized. The excitation spectrum of recombinant hmGFP has a rather wide major peak with a maximum at 490nm and a shoulder at 420nm, and its emission spectrum at 510nm. The expression of hmGFP and the chimera IPL∼hmGFP in CHO cells has shown that the fusion protein IPL∼hmGFP has retained the normal membrane targeting of the IPL from Dasyatis akajei, as well as maintaining fluorescent properties similar to those of native hmGFP, suggesting a promising prospect of the application in biotechnology research for the new protein.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 879-885 |
| Number of pages | 7 |
| Journal | Biochemical and Biophysical Research Communications |
| Volume | 301 |
| Issue number | 4 |
| DOIs | |
| State | Published - Feb 21 2003 |
| Externally published | Yes |
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Keywords
- Eukaryotic expression
- Functional cloning
- Fusion protein
- GFP homolog
- Prokaryotic expression
ASJC Scopus subject areas
- Biochemistry
- Biophysics
- Molecular Biology
Cite this
A naturally enhanced green fluorescent protein from magnificent sea anemone (Heteractis magnifica) and its functional analysis. / Tu, Hongbin; Xiong, Qian; Zhen, Suilan; Zhong, Xiaofen; Peng, Lisheng; Chen, Huiping; Jiang, Xiaoyu; Liu, Wenhua; Yang, Wenli; Wei, Jianwen; Dong, Meiling; Wu, Wenyan; Xu, Anlong.
In: Biochemical and Biophysical Research Communications, Vol. 301, No. 4, 21.02.2003, p. 879-885.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - A naturally enhanced green fluorescent protein from magnificent sea anemone (Heteractis magnifica) and its functional analysis
AU - Tu, Hongbin
AU - Xiong, Qian
AU - Zhen, Suilan
AU - Zhong, Xiaofen
AU - Peng, Lisheng
AU - Chen, Huiping
AU - Jiang, Xiaoyu
AU - Liu, Wenhua
AU - Yang, Wenli
AU - Wei, Jianwen
AU - Dong, Meiling
AU - Wu, Wenyan
AU - Xu, Anlong
PY - 2003/2/21
Y1 - 2003/2/21
N2 - A novel fluorescent protein termed hmGFP homologous to the green fluorescent protein (GFP) from Aequorea victoria was cloned from the tentacles of sea anemone Heteractis magnifica by EST sequencing and analysis of cDNA library and followed by using RT-PCR. The sequence analysis suggested that the chromophore, consensus amino acids, and secondary structure of 11 β-strands of hmGFP were similar to those of GFP from other species. The recombinant hmGFP protein with high purity was obtained by the fusion expression of pETTRX-hmGFP in Escherichia coli and subsequent purification. The pH sensitivity and fluorescence spectroscopy of recombinant hmGFP were characterized. The excitation spectrum of recombinant hmGFP has a rather wide major peak with a maximum at 490nm and a shoulder at 420nm, and its emission spectrum at 510nm. The expression of hmGFP and the chimera IPL∼hmGFP in CHO cells has shown that the fusion protein IPL∼hmGFP has retained the normal membrane targeting of the IPL from Dasyatis akajei, as well as maintaining fluorescent properties similar to those of native hmGFP, suggesting a promising prospect of the application in biotechnology research for the new protein.
AB - A novel fluorescent protein termed hmGFP homologous to the green fluorescent protein (GFP) from Aequorea victoria was cloned from the tentacles of sea anemone Heteractis magnifica by EST sequencing and analysis of cDNA library and followed by using RT-PCR. The sequence analysis suggested that the chromophore, consensus amino acids, and secondary structure of 11 β-strands of hmGFP were similar to those of GFP from other species. The recombinant hmGFP protein with high purity was obtained by the fusion expression of pETTRX-hmGFP in Escherichia coli and subsequent purification. The pH sensitivity and fluorescence spectroscopy of recombinant hmGFP were characterized. The excitation spectrum of recombinant hmGFP has a rather wide major peak with a maximum at 490nm and a shoulder at 420nm, and its emission spectrum at 510nm. The expression of hmGFP and the chimera IPL∼hmGFP in CHO cells has shown that the fusion protein IPL∼hmGFP has retained the normal membrane targeting of the IPL from Dasyatis akajei, as well as maintaining fluorescent properties similar to those of native hmGFP, suggesting a promising prospect of the application in biotechnology research for the new protein.
KW - Eukaryotic expression
KW - Functional cloning
KW - Fusion protein
KW - GFP homolog
KW - Prokaryotic expression
UR - http://www.scopus.com/inward/record.url?scp=0037458865&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0037458865&partnerID=8YFLogxK
U2 - 10.1016/S0006-291X(03)00019-6
DO - 10.1016/S0006-291X(03)00019-6
M3 - Article
C2 - 12589794
AN - SCOPUS:0037458865
VL - 301
SP - 879
EP - 885
JO - Biochemical and Biophysical Research Communications
JF - Biochemical and Biophysical Research Communications
SN - 0006-291X
IS - 4
ER -