Abstract
Contrary to the results of direct expression, various human proteins (ferritin light-chain, epithermal growth factor, interleukin-2, prepro-ghrelin, deletion mutants of glutamate decarboxylase and arginine deiminase, and mini-proinsulin) were all soluble in Escherichia coli cytoplasm when expressed with the N-terminus fusion of ferritin heavy-chain (FTN-H). Through systematic investigations, we have found that a specific peptide motif within FTN-H has a high affinity to HSP70 chaperone DnaK, and that the peptide motif was composed of a hydrophobic core of three residues (Ile, Phe and Leu) and two flanking regions enriched with polar residues (Gly, Gln and Arg). It was also observed that all the recombinant proteins expressed with the fusion of FTN-H formed spherical nanoparticles with diameters of 10-15 nm, as confirmed by the transmission electron microscopy image. The protein nanoparticles are non-covalently cross-linked supra-molecules formed by the self-assembly function of FTN-H. Upon the formation of the supra-molecule, its size is likely to be limited by the assembly properties of FTN-H, thereby keeping the self-assembled particles soluble. This study reports on the dual function of FTN-H for fusion expression and solubility enhancement of heterologous proteins: (i) high-affinity interaction with DnaK and (ii) formation of self-assembled supra-molecules with limited and constant sizes, thereby avoiding the undesirable formation of insoluble macro-aggregates of heterologous proteins.
| Original language | English |
|---|---|
| Pages (from-to) | 3751-3762 |
| Number of pages | 12 |
| Journal | Nucleic Acids Research |
| Volume | 33 |
| Issue number | 12 |
| DOIs | |
| Publication status | Published - 2005 Oct 17 |
Fingerprint
ASJC Scopus subject areas
- Genetics
Cite this
Heterologous gene expression using self-assembled supra-molecules with high affinity for HSP70 chaperone. / Ahn, Ji Young; Choi, Hyung; Kim, Yang Hoon; Han, Kyung Yeon; Park, Jin Seung; Han, Sung Sik; Lee, Jeewon.
In: Nucleic Acids Research, Vol. 33, No. 12, 17.10.2005, p. 3751-3762.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Heterologous gene expression using self-assembled supra-molecules with high affinity for HSP70 chaperone
AU - Ahn, Ji Young
AU - Choi, Hyung
AU - Kim, Yang Hoon
AU - Han, Kyung Yeon
AU - Park, Jin Seung
AU - Han, Sung Sik
AU - Lee, Jeewon
PY - 2005/10/17
Y1 - 2005/10/17
N2 - Contrary to the results of direct expression, various human proteins (ferritin light-chain, epithermal growth factor, interleukin-2, prepro-ghrelin, deletion mutants of glutamate decarboxylase and arginine deiminase, and mini-proinsulin) were all soluble in Escherichia coli cytoplasm when expressed with the N-terminus fusion of ferritin heavy-chain (FTN-H). Through systematic investigations, we have found that a specific peptide motif within FTN-H has a high affinity to HSP70 chaperone DnaK, and that the peptide motif was composed of a hydrophobic core of three residues (Ile, Phe and Leu) and two flanking regions enriched with polar residues (Gly, Gln and Arg). It was also observed that all the recombinant proteins expressed with the fusion of FTN-H formed spherical nanoparticles with diameters of 10-15 nm, as confirmed by the transmission electron microscopy image. The protein nanoparticles are non-covalently cross-linked supra-molecules formed by the self-assembly function of FTN-H. Upon the formation of the supra-molecule, its size is likely to be limited by the assembly properties of FTN-H, thereby keeping the self-assembled particles soluble. This study reports on the dual function of FTN-H for fusion expression and solubility enhancement of heterologous proteins: (i) high-affinity interaction with DnaK and (ii) formation of self-assembled supra-molecules with limited and constant sizes, thereby avoiding the undesirable formation of insoluble macro-aggregates of heterologous proteins.
AB - Contrary to the results of direct expression, various human proteins (ferritin light-chain, epithermal growth factor, interleukin-2, prepro-ghrelin, deletion mutants of glutamate decarboxylase and arginine deiminase, and mini-proinsulin) were all soluble in Escherichia coli cytoplasm when expressed with the N-terminus fusion of ferritin heavy-chain (FTN-H). Through systematic investigations, we have found that a specific peptide motif within FTN-H has a high affinity to HSP70 chaperone DnaK, and that the peptide motif was composed of a hydrophobic core of three residues (Ile, Phe and Leu) and two flanking regions enriched with polar residues (Gly, Gln and Arg). It was also observed that all the recombinant proteins expressed with the fusion of FTN-H formed spherical nanoparticles with diameters of 10-15 nm, as confirmed by the transmission electron microscopy image. The protein nanoparticles are non-covalently cross-linked supra-molecules formed by the self-assembly function of FTN-H. Upon the formation of the supra-molecule, its size is likely to be limited by the assembly properties of FTN-H, thereby keeping the self-assembled particles soluble. This study reports on the dual function of FTN-H for fusion expression and solubility enhancement of heterologous proteins: (i) high-affinity interaction with DnaK and (ii) formation of self-assembled supra-molecules with limited and constant sizes, thereby avoiding the undesirable formation of insoluble macro-aggregates of heterologous proteins.
UR - http://www.scopus.com/inward/record.url?scp=22244436285&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=22244436285&partnerID=8YFLogxK
U2 - 10.1093/nar/gki692
DO - 10.1093/nar/gki692
M3 - Article
VL - 33
SP - 3751
EP - 3762
JO - The BMJ
JF - The BMJ
SN - 0730-6512
IS - 12
ER -