CYSTEINE-SPECIFIC SURFACE TETHERING OF GENETICALLY-ENGINEERED CYTOCHROMES FOR FABRICATION OF METALLOPROTEIN NANOSTRUCTURES
CYSTEINE-SPECIFIC SURFACE TETHERING OF GENETICALLY-ENGINEERED CYTOCHROMES FOR FABRICATION OF METALLOPROTEIN NANOSTRUCTURES
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DOI：
10.1021/la00013a023
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发表时间：
1994-01-01
期刊：
影响因子：
3.9
通讯作者：
BOHN, PW
中科院分区：
文献类型：
--
作者：
HONG, HG;JIANG, M;BOHN, PW
通过在固体和蛋白质表面上引入特定的和互补的反应基团来制备定向的金属蛋白质纳米结构，这在很大程度上取决于用于制备固体表面的反应条件。关键问题包括Si-O键的水解稳定性，简单亲核硅烷试剂的低反应性，蛋白质物理吸附，以及确定产生单层蛋白质覆盖的条件。这些问题在很大程度上可以通过利用两步连接体合成来避免，其中首先用单层（3-氨丙基）硅烷（S-APS）制备表面，然后用异双功能试剂N-琥珀酰亚胺基6-马来酰亚胺基己酸酯（EMCS）衍生化所得结构。然后将马来酰亚胺官能团呈递给蛋白质，其中通过基因工程技术引入了单个独特的半胱氨酸残基。水解稳定性通过包括反应后固化步骤而显著增强，其中固体表面温度升高以驱动烷氧基硅烷缩合反应完成。最后，用气相化学气相沉积程序代替3-APS的液相反应，可以显著更好地控制所得薄膜的覆盖率和质量。
The preparation of oriented metalloprotein nanostructures through introduction of specific and complementary reactive groups on the solid and protein surfaces is critically dependent on the reaction conditions used to prepare the solid surface. Key problems include the hydrolytic stability of the Si-O bond, the low reactivity of simple nucleophilic silane reagents, protein physisorption, and identification of conditions for producing monolayer protein coverages. These problems are largely circumvented by utilizing a two-step linker synthesis, in which the surface is first prepared with a monolayer of (3-aminopropyl)silane (S-APS), and the resulting structure is derivatized with the heterobifunctional reagent N-succinimidyl 6-maleimidocaproate (EMCS). The maleimide functionality is then presented to the protein, into which a single unique cysteine residue has been introduced by genetic engineering techniques. Hydrolytic stability is dramatically enhanced by including a postreaction curing step, in which the solid surface temperature is elevated to drive the alkoxysilane condensation reaction to completion. Finally substituing a gas-phase chemical vapor deposition procedure for the liquid-phase reaction of the 3-APS produces dramatically better control over coverage and quality of the resulting films.