Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Pawińskiego 5a, 02-106 Warsaw, Poland.
Inorg Chem. 2024 Jul 1;63(26):12323-12332. doi: 10.1021/acs.inorgchem.4c01797. Epub 2024 Jun 13.
The choice of correct pH buffer is crucial in chemical studies modeling biological processes involving Cu ions. Popular buffers for physiological pH are known to form Cu(II) complexes, but their impact on kinetics of Cu(II) complexation has not been considered. We performed a stopped-flow kinetic study of Cu ion interactions with four popular buffers (phosphate, Tris, HEPES, and MOPS) and two buffers considered as nonbinding (MES and PIPPS). Next, we studied their effects on the rate of Cu reaction with Gly-Gly-His (GGH), a tripeptide modeling physiological Cu(II) sites, which we studied previously at conditions presumably excluding the buffer interference [Kotuniak, R.; 2020, 59, 11234-11239]. We observed that (i) all tested pH 7.4 buffers formed Cu(II) complexes within the stopped-flow instrument dead time; (ii) Cu(II)-peptide complexes were formed via ternary complexes with the buffers; (iii) nevertheless, Good buffers affected the observed rate of Cu(II)-GGH complex formation only slightly; (iv) Tris was a competitive inhibitor of Cu(II)-GGH complexation; while (v) phosphate was a reaction catalyst. This is particularly important as phosphate is a biological buffer.
在模拟涉及 Cu 离子的生物过程的化学研究中，正确 pH 缓冲液的选择至关重要。用于生理 pH 的常用缓冲液已知会形成 Cu(II)配合物，但尚未考虑它们对 Cu(II)配合物动力学的影响。我们进行了一项 Cu 离子与四种常用缓冲液（磷酸盐、Tris、HEPES 和 MOPS）和两种被认为不结合的缓冲液（MES 和 PIPPS）相互作用的停流动力学研究。然后，我们研究了它们对 Cu 与 Gly-Gly-His（GGH）反应速率的影响，GGH 是一种模拟生理 Cu(II)位点的三肽，我们之前在可能排除缓冲干扰的条件下研究了该反应[Kotuniak, R.; 2020, 59, 11234-11239]。我们观察到：(i) 所有测试的 pH 7.4 缓冲液在停流仪器死时间内形成 Cu(II)配合物；(ii) Cu(II)-肽配合物通过与缓冲液的三元配合物形成；(iii) 然而，Good 缓冲液仅对观察到的 Cu(II)-GGH 配合物形成速率产生轻微影响；(iv) Tris 是 Cu(II)-GGH 配合物形成的竞争性抑制剂；而 (v) 磷酸盐是反应催化剂。这一点尤为重要，因为磷酸盐是一种生物缓冲液。
