Synthesis and conformational preferences of peptides and proteins with cysteine sulfonic acid
Synthesis and conformational preferences of peptides and proteins with cysteine sulfonic acid
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DOI：
10.1039/d3ob00179b
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发表时间：
2023-03-07
影响因子：
3.2
通讯作者：
Zondlo，Neal J.
中科院分区：
文献类型：
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作者：
Bhatt，Megh R.;Zondlo，Neal J.
半胱氨酸磺酸(Cys-SO3H)是半胱氨酸的氧化翻译后修饰，由半胱氨酸亚磺酸进一步氧化而成。半胱氨酸磺酸被认为是一种不可逆的翻译后修饰，它是导致蛋白质氧化损伤的氧化应激的生物标志物。半胱氨酸磺酸是一种阴离子的磺酸盐(Cys-SO3−；半胱酸盐)，其电离状态几乎只存在于生理pH(pKa∼−2)。为了了解半胱氨酸磺酸氧化过程中蛋白质结构的变化，我们利用实验方法、生物信息学和基于密度泛函理论的计算分析方法，分析了其构象偏好。将半胱氨酸磺酸引入到α-螺旋和多聚脯氨酸II螺旋的模型肽中。在多肽中，在室温下，半胱氨酸在甲基三氧基苯(MeReO_3)和H_2O_2的溶液中快速而有效地氧化为磺酸。以三苯基保护的半胱氨酸为原料，用MeREO_3和H_2O_2氧化，在固相上生成了含有半胱氨酸磺酸的多肽。利用甲氧基苄基(MOB)保护的半胱氨酸，用MeREO_3和H_2O_2固相氧化，在完全合成的多肽中生成了半胱氨酸-硫代半胱氨酸(−)的MOB前驱体。这两种固相法能够以实际的方式合成含有Cys-SO3−或Cys-SO2−的多肽，而不需要溶液相合成。Cys-SO3−具有较低的PPII繁殖倾向，尽管促进了ϕ中相对紧密的构象。相反，在PPII启动模型体系中，Cys-SO3−相对中性Cys促进PPII，PPII启动类似于Cys硫代，但低于Cys-SO2−或ALA。在α-螺旋模型体系中，−通过磺酸盐-酰胺侧链-主链氢键的有利螺旋偶极相互作用和有利的α-螺旋封端，促进了N-末端附近的α-螺旋。在所有的多肽中，磺酸盐侧链的有序性明显低于亚磺酸盐。对PDB中Cys-SO_3−的分析表明，Cys-SO_3−有很强的局部(I/I或I/I+1)侧链-主链磺酸盐-酰胺氢键的倾向，80%的Cys-SO_3−残基表现出这种相互作用。密度泛函理论计算表明，磺酸盐与残基内酰胺和/或I+1酰胺的侧链-主链氢键是有利的。然而，由于磺酸盐中氧的电荷密度较低，磺酸盐与水或酰胺的氢键以及与亲氧金属的相互作用比亚磺酸盐弱。
Cysteine sulfonic acid (Cys-SO3H; cysteic acid) is an oxidative post-translational modification of cysteine, resulting from further oxidation from cysteine sulfinic acid (Cys-SO2H). Cysteine sulfonic acid is considered an irreversible post-translational modification, which serves as a biomarker of oxidative stress that has resulted in oxidative damage to proteins. Cysteine sulfonic acid is anionic, as a sulfonate (Cys-SO3−; cysteate), in the ionization state that is almost exclusively present at physiological pH (pKa ∼ −2). In order to understand protein structural changes that can occur upon oxidation to cysteine sulfonic acid, we analyzed its conformational preferences, using experimental methods, bioinformatics, and DFT-based computational analysis. Cysteine sulfonic acid was incorporated into model peptides for α-helix and polyproline II helix (PPII). Within peptides, oxidation of cysteine to the sulfonic acid proceeds rapidly and efficiently at room temperature in solution with methyltrioxorhenium (MeReO3) and H2O2. Peptides containing cysteine sulfonic acid were also generated on solid phase using trityl-protected cysteine and oxidation with MeReO3 and H2O2. Using methoxybenzyl (Mob)-protected cysteine, solid-phase oxidation with MeReO3 and H2O2 generated the Mob sulfone precursor to Cys-SO2− within fully synthesized peptides. These two solid-phase methods allow the synthesis of peptides containing either Cys-SO3− or Cys-SO2− in a practical manner, with no solution-phase synthesis required. Cys-SO3− had low PPII propensity for PPII propagation, despite promoting a relatively compact conformation in ϕ. In contrast, in a PPII initiation model system, Cys-SO3− promoted PPII relative to neutral Cys, with PPII initiation similar to Cys thiolate but less than Cys-SO2− or Ala. In an α-helix model system, Cys-SO3− promoted α-helix near the N-terminus, due to favorable helix dipole interactions and favorable α-helix capping via a sulfonate-amide side chain–main chain hydrogen bond. Across all peptides, the sulfonate side chain was significantly less ordered than that of the sulfinate. Analysis of Cys-SO3− in the PDB revealed a very strong propensity for local (i/i or i/i + 1) side chain–main chain sulfonate–amide hydrogen bonds for Cys-SO3−, with >80% of Cys-SO3− residues exhibiting these interactions. DFT calculations conducted to explore these conformational preferences indicated that side chain–main chain hydrogen bonds of the sulfonate with the intraresidue amide and/or with the i + 1 amide were favorable. However, hydrogen bonds to water or to amides, as well as interactions with oxophilic metals, were weaker for the sulfonate than the sulfinate, due to lower charge density on the oxygens in the sulfonate.