College of Chemistry and Chemical Engineering, Shenyang Normal University, Shenyang 110034, China.
College of Chemistry, Liaoning University, Shenyang, 110036, China.
Analyst. 2024 Sep 23;149(19):4960-4969. doi: 10.1039/d4an00421c.
The effect of anions on the positive electrospray ionization (ESI) of proteins in different strong acids with varying pH values from 3 to 1 is studied using high-pressure ESI. Reducing the pH from ∼2 to 1 caused a drastic shift in charge state from a high-charge-state distribution (HCSD) to a narrow low-charge-state distribution (LCSD). The shift in charge state was consistent with the circular dichroism result that showed a conformational change due to the "acid-induced folding" of proteins from an unfolding state to a compact molten globule state. Acids of different anions produced noticeable differences in the average charge for HCSD and LCSD. For HCSD, the average charge was lower than the value typically observed using formic and acetic acids. As for LCSD, the average charge was lower than the "native" charge. The high abundance of acid anion that induces the protein compaction was believed to play a role in charge reduction. The effectiveness of anions to "" a highly unfolded protein to a compact state and the propensity to reduce the charge of HCSD for proteins appeared to follow the selectivity series of anions towards the stationary phase in ion chromatography. However, the propensity of anions to reduce the charge for LCSD follows quite an opposite trend. The presence of ammonium salt in the acidic solution was found to increase the charge of LCSD. The simple mass spectrum with a narrow distribution of charge state obtained with perchloric acid at pH 1 was demonstrated to facilitate the counting of basic sites.
采用高压电喷雾技术研究了在不同 pH 值（3 至 1）的强酸中，阴离子对蛋白质正电喷雾电离（ESI）的影响。将 pH 值从约 2 降低至 1 会导致电荷状态从高电荷态分布（HCSD）急剧转变为窄低电荷态分布（LCSD）。电荷状态的转变与圆二色性结果一致，表明由于“酸诱导折叠”，蛋白质从展开状态转变为紧凑的无规卷曲状态，从而发生构象变化。不同阴离子的酸在 HCSD 和 LCSD 的平均电荷上产生了明显的差异。对于 HCSD，平均电荷低于使用甲酸和乙酸通常观察到的值。对于 LCSD，平均电荷低于“天然”电荷。据信，诱导蛋白质紧密化的高丰度酸阴离子在电荷减少中起作用。阴离子将高度展开的蛋白质“折叠”成紧凑状态的有效性以及降低 HCSD 蛋白质电荷的倾向似乎遵循了离子色谱中阴离子对固定相的选择性系列。然而，阴离子降低 LCSD 电荷的倾向则遵循完全相反的趋势。在酸性溶液中存在铵盐时，发现 LCSD 的电荷增加。在 pH 值为 1 时使用高氯酸获得的具有窄电荷态分布的简单质谱被证明有利于基本位点的计数。
