Seker Urartu O S, Wilson Brandon, Sahin Deniz, Tamerler Candan, Sarikaya Mehmet
Materials Science and Engineering, University of Washington, Seattle, Washington 98195, USA.
Biomacromolecules. 2009 Feb 9;10(2):250-7. doi: 10.1021/bm8009895.
Binding kinetics of platinum-, silica-, and gold-binding peptides were investigated using a modified surface plasmon resonance spectroscopy (SPR). Platinum binding septa-peptides, quartz-binding dodecapeptides, and gold-binding 14-aa peptides were originally selected using phage or cell surface display libraries using the mineral or pure forms of these materials. All of the peptides were synthesized singly to investigate their binding kinetics and to assess quantitatively the specific affinity of each to its material of selection. The peptides were also postselection engineered to contain multiple copies of the same original sequences to quantify the effects of repeating units. SPR spectroscopy, normally using gold surfaces, was modified to contain a thin film (a few nm thick) of the material of interest (silica or platinum) on gold to allow the quantitative study of the adsorption kinetics of specific solid-binding peptides. The SPR experiments, carried out at different concentrations, on all three materials substrates, resulted in Langmuir behavior that allowed the determination of the kinetic parameters, including adsorption, desorption, and equilibrium binding constants for each of the solids as well as free energy of adsorption. Furthermore, we also tested multiple repeats of the peptide sequences, specifically three repeats, to see if there is a general trend of increased binding with increased number of binding domains. There was no general trend in the binding strength of the peptides with the increase of the repeat units from one to three, possibly because of the conformational changes between the single and multiple repeat polypeptides. In all cases, however, the binding was strong enough to suggest that these inorganic binding peptides could potentially be used as specific molecular linkers to bind molecular entities to specific solid substrates due to their surface recognition characteristics.
使用改良的表面等离子体共振光谱法(SPR)研究了铂结合肽、二氧化硅结合肽和金结合肽的结合动力学。铂结合七肽、石英结合十二肽和金结合14氨基酸肽最初是使用噬菌体或细胞表面展示文库,通过这些材料的矿物形式或纯形式筛选出来的。所有肽均单独合成,以研究其结合动力学,并定量评估每种肽对其选择材料的特异性亲和力。这些肽还经过筛选后工程改造,包含多个相同原始序列的拷贝,以量化重复单元的影响。通常使用金表面的SPR光谱法经过改良,在金表面含有一层感兴趣材料(二氧化硅或铂)的薄膜(几纳米厚),以便对特定固体结合肽的吸附动力学进行定量研究。在所有三种材料底物上,于不同浓度下进行的SPR实验呈现出朗缪尔行为,从而能够确定动力学参数,包括每种固体的吸附、解吸和平衡结合常数以及吸附自由能。此外,我们还测试了肽序列的多个重复,特别是三个重复,以查看是否存在随着结合域数量增加而结合增强的一般趋势。随着重复单元从一个增加到三个,肽的结合强度没有一般趋势,这可能是由于单重复和多重复多肽之间的构象变化。然而,在所有情况下,结合都足够强,表明这些无机结合肽由于其表面识别特性,有可能用作特定分子连接体,将分子实体与特定固体底物结合。