Chemistry Department, The University of Western Ontario, London, Ontario, Canada.
Chemistry. 2011 Jun 6;17(24):6744-52. doi: 10.1002/chem.201003535. Epub 2011 May 3.
The synthesis and electrochemical properties of ferrocene conjugates are presented for the purpose of investigating adenosine 5'-[γ-ferrocenoylalkyl] triphosphate (1 a-4 a, ferrocene (Fc)-ATP) as co-substrates for phosphorylation reactions. Compounds 1 a-4 a were synthesized, purified by HPLC, and characterized by NMR spectroscopy and mass spectrometry. In solution, all Fc-ATP bioconjugates exhibit a reversible one-electron redox process with a half-wave potential (E(1/2)) in the 390-430 mV range, peak separations (ΔE(p)) in the 40-70 mV range, and the peak current ratio (i(pa)/i(pc)) near unity. The peptide-modified surface Glu-Gly-Ile-Tyr-Asp-Val-Pro was used to study the sarcoma-related protein (Src) kinase activity by employing the Fc-ATP bioconjugates as co-substrates. Subsequent kinase-catalyzed transfer of the γ-Fc-phosphate group to the tyrosine residues of the surface-bound peptides was characterized by a formal potential (E°) ≈390 mV (vs. Ag/AgCl). The Fc-coverage, estimated by time-of-flight secondary-ion mass spectrometry (TOF-SIMS) and cyclic voltammetry (CV), suggested validity of Fc-ATP conjugates as kinase co-substrates. Depending on the length of the alkyl spacer of the Fc-ATP conjugate, different current densities were obtained, pointing to a direct correlation between the two. Molecular modeling revealed that the structural constraint imposed by the short alkyl spacer (1 a) causes a steric congestion and negatively affects the outcome of phosphorylation reaction. An optimal analytical response was obtained with the Fc-ATP conjugates with linker lengths longer than six CH(2) groups.
本文介绍了二茂铁衍生物的合成及电化学性质，旨在研究腺苷 5'-[γ-二茂铁酰基]三磷酸酯（1a-4a，二茂铁（Fc）-ATP）作为磷酸化反应的共底物。合成了化合物 1a-4a，通过 HPLC 进行纯化，并通过 NMR 光谱和质谱进行了表征。在溶液中，所有 Fc-ATP 生物缀合物均表现出可逆的单电子氧化还原过程，半波电位（E(1/2)）在 390-430 mV 范围内，峰分离（ΔE(p)）在 40-70 mV 范围内，峰电流比（i(pa)/i(pc)）接近 1。使用表面 Glu-Gly-Ile-Tyr-Asp-Val-Pro 肽修饰，将 Fc-ATP 生物缀合物用作共底物，研究肉瘤相关蛋白（Src）激酶活性。随后通过激酶催化将 γ-Fc-磷酸基团转移到表面结合肽的酪氨酸残基上，其形式电位（E°）≈390 mV（相对于 Ag/AgCl）。通过飞行时间二次离子质谱（TOF-SIMS）和循环伏安法（CV）估计的 Fc 覆盖率表明 Fc-ATP 缀合物作为激酶共底物是有效的。根据 Fc-ATP 缀合物的烷基间隔物的长度，获得了不同的电流密度，这表明两者之间存在直接相关性。分子建模表明，短烷基间隔物（1a）施加的结构约束会导致空间位阻，从而对磷酸化反应的结果产生负面影响。对于具有长于六个 CH(2)基团的连接物长度的 Fc-ATP 缀合物，获得了最佳的分析响应。
