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
C2 - 20143369
AN - SCOPUS:77950827616
VL - 5
SP - 877
EP - 886
JO - Chemistry - An Asian Journal
JF - Chemistry - An Asian Journal
SN - 1861-4728
IS - 4
ER -