State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210093, China.
Phys Chem Chem Phys. 2012 Jul 14;14(26):9460-7. doi: 10.1039/c2cp40594f. Epub 2012 May 31.
A nanochannel array based nanodevice can mimic the biological environments and thus unveil the natural properties, conformation and recognition information of biomolecules such as proteins and DNA in confined spaces. Here we report that porous anodic alumina (PAA) of a highly parallel nanochannel array covalently modified with proteins significantly modulates the transport of a negatively charged probe of ferricyanide due to the electrostatic interactions between the probes and modified nanochannel inner surface. Results show that such electrostatic interaction exists in a wide range of ionic strength from 1 mM to 100 mM in 20 nm nanochannels modified with proteins (hemoglobin, bovine serum albumin, and goat anti-rabbit IgG secondary antibody). In addition, the maximal steady-state flux of the charged probe through the modified nanochannel array is directly related to the ionic strength which determines the electric double layer thickness and solution pH which modulates the nanochannel surface charge. Thus, the modulated mass transport of the probe by solution pH can be used to study the charge properties of the immobilized proteins in nanochannel confined conditions, leading us to obtain the isoelectric point (pI) of the proteins confined in nanochannels. The determined pI values of two known proteins of hemoglobin and bovine serum albumin are close to the ones of the same proteins covalently modified on a 3-mercaptopropionic acid self-assembled monolayer/gold electrode. In addition, the pI of an unknown protein of goat anti-rabbit IgG secondary antibody confined in nanochannels was determined to be 6.3. Finally, the confinement effect of nanochannels on the charge properties of immobilized proteins has been discussed.
基于纳米通道阵列的纳米器件可以模拟生物环境，从而揭示蛋白质和 DNA 等生物分子在受限空间中的天然性质、构象和识别信息。在这里，我们报告说，高度平行的纳米通道阵列的多孔阳极氧化铝（PAA）通过共价键与蛋白质修饰，可以显著调节带负电荷的铁氰化物探针的传输，这是由于探针和修饰的纳米通道内表面之间的静电相互作用。结果表明，这种静电相互作用存在于广泛的离子强度范围内，从 1mM 到 100mM，在 20nm 纳米通道中用蛋白质（血红蛋白、牛血清白蛋白和兔抗山羊 IgG 二抗）修饰。此外，带电探针通过修饰的纳米通道阵列的最大稳态通量与离子强度直接相关，离子强度决定了双电层厚度和溶液 pH 值，从而调节纳米通道表面电荷。因此，通过溶液 pH 值调制探针的传质可以用于研究在纳米通道受限条件下固定化蛋白质的电荷特性，使我们能够获得在纳米通道中受限的蛋白质的等电点（pI）。两种已知蛋白质血红蛋白和牛血清白蛋白的确定 pI 值接近在 3-巯基丙酸自组装单层/金电极上共价修饰的相同蛋白质的 pI 值。此外，还确定了在纳米通道中受限的兔抗山羊 IgG 二抗未知蛋白质的 pI 值为 6.3。最后，讨论了纳米通道对固定化蛋白质电荷特性的限制作用。
