Real‐time analysis of the calcium‐dependent interaction between calmodulin and a synthetic oligopeptide of calcineurin by a surface plasmon resonance biosensor
Real‐time analysis of the calcium‐dependent interaction between calmodulin and a synthetic oligopeptide of calcineurin by a surface plasmon resonance biosensor
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通过表面等离子共振生物传感器实时分析钙调蛋白和钙调磷酸酶合成寡肽之间的钙依赖性相互作用
DOI：
10.1016/0014-5793(94)00965-1
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发表时间：
1994
期刊：
影响因子：
3.5
通讯作者：
M. Maki
中科院分区：
文献类型：
--
作者：
E. Takano;Masakazu Hatanaka;M. Maki
使用自动表面等离子体共振生物传感器BIAcore分析钙调蛋白（CaM）与人钙调磷酸酶A-2的预测CaM结合区域的合成寡肽之间的钙依赖性相互作用。通过巯基二硫化物交换法，将寡肽经由氨基末端半胱氨酸残基固定到生物传感器芯片上。每次分析后，通过含EGTA的缓冲液再生生物传感器芯片。动力学实验表明，钙调素结合与高亲和力的寡肽在钙离子依赖性的方式。结合速率常数kass和解离速率常数kdiss分别为2.3 × 10 - 5 M-1·s-1和3.9 × 10 - 3s-1。kdiss/kass的比值为1.7 × 10− 8 M，与平衡相测定的解离常数（Kd）2.4 × 10− 8 M非常一致。
The calcium‐dependent interaction between calmodulin (CaM) and the synthetic oligopeptide of a predicted CaM‐binding region of human calcineurin A‐2 was analysed with an automated surface plasmon resonance biosensor, BIAcore. The oligopeptide was immobilized to a biosensor chip via the amino‐terminal cysteine residue by a thioldisulphide exchange method. The biosensor chip was regenerated by an EGTA‐containing buffer after each analysis. Kinetics experiments showed that CaM bound with a high affinity to the oligopeptide in a Ca2+‐dependent manner. The estimated rate constants of association (kass) and dissociation (kdiss) were 2.3 × 1O5M−1·s−1and 3.9 × 10−3s−1, respectively. The ratio ofkdiss/kass, 1.7 × 10−8M, was in good agreement with the dissociation constant (Kd) of 2.4 × 10−8M determined from the equilibrium phase.