Abstract
A complementary recognition pair of a short-peptide tag and a small molecular probe is a versatile molecular tool for protein detection, handling, and purification, and so forth. In this manuscript, we report that the binuclear Ni II -DpaTyr (DpaTyr = bis((dipicolylamino)methyl)-tyrosine) complex serves as a strong binding probe for an oligo-aspartate tag tethered to a protein. Among various binuclear metal complexes of M-DpaTyr (M = Zn II , Ni II , Mn II , Cu II , Cd II , Co III , and Fe III ), we have found that Ni II -DpaTyr (1-2Ni II ) displays a strongbinding affinity (apparent binding constant: K app ≈10 5 M -1 ) for an oligo-aspartate peptide under neutral aqueous conditions (50 mM HEPES, 100 mM NaCl, pH 7.2). Detailed isothermal-titration calorimetry (ITC) studies reveal that the tri-aspartate D3-tag (DDD) is an optimal sequence recognized by 1-2Ni II in a 1:1 binding stoichiometry. On the other hand, other metal complexes of DpaTyr, except for Ni II -and Zn II - DpaTyr, show a negligible binding affinity for the oligo-aspartate peptide. The binding affinity was greatly enhanced in the pair between the dimer of Ni II -DpaTyr and the repeated D3 tag peptide (D3x2), such as DDDXXDDD, on the basis of the multivalent coordination interaction between them. Most notably, a remarkably high-binding affinity (K app 10 9 M -1 ) was achieved between the Ni II -DpaTyr dimer 4-4Ni II and the D3 x 2 tag peptide (DDDNGDDD). This affinity is ≈fold stronger than that observed in the binding pair of the Zn II -DpaTyr (4-4Zn II ) and the D4x2 tag (DDDDGDDDD), a useful tagprobe pair previously reported by us. The recognition pair of the Ni II -DpaTyr probe and the D3 x 2 tag can also work effectively on a protein surface, that is, 4-4Ni II is strongly bound to the FKBP12 protein tethered with the D3 x 2 tag (DDDNGDDD) with a large K app value of 5 x 10 8 m -1 . Taking advantage of the strong-binding affinity, this pair was successfully applied to the selective inactivation of the tagfused (β-galactosidase by using the chromophore-assisted light inactivation (CALI) technique under crude conditions, such as cell lysate.
| Original language | English |
|---|---|
| Pages (from-to) | 877-886 |
| Number of pages | 10 |
| Journal | Chemistry - An Asian Journal |
| Volume | 5 |
| Issue number | 4 |
| DOIs | |
| Publication status | Published - Apr 1 2010 |
| Externally published | Yes |
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All Science Journal Classification (ASJC) codes
- Biochemistry
- Organic Chemistry
Cite this
Binuclear Ni II -DpaTyr complex as a high affinity probe for an oligo-aspartate tag tethered to proteins . / Ojida, Akio; Fujishima, Sho Hei; Honda, Kei; Nonaka, Hiroshi; Uchinomiya, Shohei; Hamachi, Itaru.
In: Chemistry - An Asian Journal, Vol. 5, No. 4, 01.04.2010, p. 877-886.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Binuclear Ni II -DpaTyr complex as a high affinity probe for an oligo-aspartate tag tethered to proteins
AU - Ojida, Akio
AU - Fujishima, Sho Hei
AU - Honda, Kei
AU - Nonaka, Hiroshi
AU - Uchinomiya, Shohei
AU - Hamachi, Itaru
PY - 2010/4/1
Y1 - 2010/4/1
N2 - A complementary recognition pair of a short-peptide tag and a small molecular probe is a versatile molecular tool for protein detection, handling, and purification, and so forth. In this manuscript, we report that the binuclear Ni II -DpaTyr (DpaTyr = bis((dipicolylamino)methyl)-tyrosine) complex serves as a strong binding probe for an oligo-aspartate tag tethered to a protein. Among various binuclear metal complexes of M-DpaTyr (M = Zn II , Ni II , Mn II , Cu II , Cd II , Co III , and Fe III ), we have found that Ni II -DpaTyr (1-2Ni II ) displays a strongbinding affinity (apparent binding constant: K app ≈10 5 M -1 ) for an oligo-aspartate peptide under neutral aqueous conditions (50 mM HEPES, 100 mM NaCl, pH 7.2). Detailed isothermal-titration calorimetry (ITC) studies reveal that the tri-aspartate D3-tag (DDD) is an optimal sequence recognized by 1-2Ni II in a 1:1 binding stoichiometry. On the other hand, other metal complexes of DpaTyr, except for Ni II -and Zn II - DpaTyr, show a negligible binding affinity for the oligo-aspartate peptide. The binding affinity was greatly enhanced in the pair between the dimer of Ni II -DpaTyr and the repeated D3 tag peptide (D3x2), such as DDDXXDDD, on the basis of the multivalent coordination interaction between them. Most notably, a remarkably high-binding affinity (K app 10 9 M -1 ) was achieved between the Ni II -DpaTyr dimer 4-4Ni II and the D3 x 2 tag peptide (DDDNGDDD). This affinity is ≈fold stronger than that observed in the binding pair of the Zn II -DpaTyr (4-4Zn II ) and the D4x2 tag (DDDDGDDDD), a useful tagprobe pair previously reported by us. The recognition pair of the Ni II -DpaTyr probe and the D3 x 2 tag can also work effectively on a protein surface, that is, 4-4Ni II is strongly bound to the FKBP12 protein tethered with the D3 x 2 tag (DDDNGDDD) with a large K app value of 5 x 10 8 m -1 . Taking advantage of the strong-binding affinity, this pair was successfully applied to the selective inactivation of the tagfused (β-galactosidase by using the chromophore-assisted light inactivation (CALI) technique under crude conditions, such as cell lysate.
AB - A complementary recognition pair of a short-peptide tag and a small molecular probe is a versatile molecular tool for protein detection, handling, and purification, and so forth. In this manuscript, we report that the binuclear Ni II -DpaTyr (DpaTyr = bis((dipicolylamino)methyl)-tyrosine) complex serves as a strong binding probe for an oligo-aspartate tag tethered to a protein. Among various binuclear metal complexes of M-DpaTyr (M = Zn II , Ni II , Mn II , Cu II , Cd II , Co III , and Fe III ), we have found that Ni II -DpaTyr (1-2Ni II ) displays a strongbinding affinity (apparent binding constant: K app ≈10 5 M -1 ) for an oligo-aspartate peptide under neutral aqueous conditions (50 mM HEPES, 100 mM NaCl, pH 7.2). Detailed isothermal-titration calorimetry (ITC) studies reveal that the tri-aspartate D3-tag (DDD) is an optimal sequence recognized by 1-2Ni II in a 1:1 binding stoichiometry. On the other hand, other metal complexes of DpaTyr, except for Ni II -and Zn II - DpaTyr, show a negligible binding affinity for the oligo-aspartate peptide. The binding affinity was greatly enhanced in the pair between the dimer of Ni II -DpaTyr and the repeated D3 tag peptide (D3x2), such as DDDXXDDD, on the basis of the multivalent coordination interaction between them. Most notably, a remarkably high-binding affinity (K app 10 9 M -1 ) was achieved between the Ni II -DpaTyr dimer 4-4Ni II and the D3 x 2 tag peptide (DDDNGDDD). This affinity is ≈fold stronger than that observed in the binding pair of the Zn II -DpaTyr (4-4Zn II ) and the D4x2 tag (DDDDGDDDD), a useful tagprobe pair previously reported by us. The recognition pair of the Ni II -DpaTyr probe and the D3 x 2 tag can also work effectively on a protein surface, that is, 4-4Ni II is strongly bound to the FKBP12 protein tethered with the D3 x 2 tag (DDDNGDDD) with a large K app value of 5 x 10 8 m -1 . Taking advantage of the strong-binding affinity, this pair was successfully applied to the selective inactivation of the tagfused (β-galactosidase by using the chromophore-assisted light inactivation (CALI) technique under crude conditions, such as cell lysate.
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U2 - 10.1002/asia.200900362
DO - 10.1002/asia.200900362
M3 - Article
VL - 5
SP - 877
EP - 886
JO - Chemistry - An Asian Journal
JF - Chemistry - An Asian Journal
SN - 1861-4728
IS - 4
ER -