Aromatic side chain-porphyrin interactions in designed hemoproteins
Aromatic side chain-porphyrin interactions in designed hemoproteins
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DOI：
10.1021/ja990606r
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发表时间：
1999-12-22
影响因子：
15
通讯作者：
Benson, DR
中科院分区：
文献类型：
--
作者：
Liu, DH;Williamson, DA;Benson, DR
通常观察到芳香族氨基酸侧链与天然血红素蛋白的血红素辅因子相互作用。这些相互作用是先前针对蛋白质中的芳香族氨基酸侧链对或组所确定的类型：偏移π堆积和T堆积（边到面排列）。为了评估这种相互作用如何影响血红素蛋白的结构稳定性，我们合成了肽夹心中血红素（PSM）2和3，其中1中的丙氨酸-4（Ala-4）残基已分别被苯丙氨酸（Phe）和色氨酸（Trp）取代。还制备了1、2和3的Co（III）类似物（分别为1-Co、2-Co和3-Co）。1中的组氨酸（His）与铁的配位先前已显示出诱导肽中的螺旋构象（在8 ℃下螺旋含量类似于50%）。分子模拟研究表明，色氨酸，而不是苯丙氨酸，可以从事边到面的相互作用与卟啉，如果肽是完全螺旋。因此，预测用Trp替代Ala-4而不是用Phe替代Ala-4有利于增强肽螺旋含量。圆二色光谱与3中相对于i显著增加的螺旋含量一致，但在2中不一致。然而，通过电喷雾电离质谱法测定的氢-氘（H/D）交换速率以远大于2 > 3的顺序降低，而pH滴定揭示模型蛋白质折叠的稳定性以远大于1的顺序3 > 2降低。此外，2-Co和3-Co的H-1 NMR谱表明，在每个化合物中的芳族侧链的卟啉环的屏蔽区域内取向。二维NOE和化学位移数据表明，3-Co中的螺旋比1-Co中的螺旋更有组织，并且跨越几乎整个肽序列，而在2-Co中，中间稳定性的较短螺旋在Phe-4和Ala-13之间运行。结合的结果表明，芳香族侧链卟啉相互作用2和3稳定各自的模型蛋白质折叠，并建议在天然血红素蛋白的相应的相互作用类似的作用。最后，在3-Co中的色氨酸侧链的化学位移模式，苯丙氨酸和色氨酸对肽结构的不同影响，以及所有三个Co（III）PSM中的α-NH和α-CH氢所表现出的化学位移模式表明，这些设计的血红素蛋白的溶液结构与分子建模研究中预测的那些相似。
Aromatic amino acid side chains are commonly observed to interact with the heme cofactors of natural hemoproteins. These interactions are of the types previously identified for pairs or groups of aromatic amino acid side chains in proteins: offset pi-stacking and T-stacking (an edge-to-face arrangement). To evaluate how such interactions may influence structural stability of hemoproteins, we synthesized peptide-sandwiched mesohemes (PSMs) 2 and 3 in which the alanine-4 (Ala-4) residues in 1 have been replaced by phenylalanine (Phe) and tryptophan (Trp), respectively. The Co(III) analogues of 1, 2, and 3 (l-Co, 2-Co, and 3-Co, respectively) were also prepared. Histidine (His)-to-iron coordination in 1 had previously been shown to induce helical conformations in the peptides (helix content similar to 50% at 8 degrees C). Molecular modeling studies suggested that Trp, but not Phe, could engage in edge-to-face interactions with the porphyrin if the peptides are fully helical. Replacing Ala-4 with Trp, bur not with Phe, was thus predicted to Favor enhanced peptide helix content. Circular dichroism spectra are consistent with significantly increased helix content in 3 relative to i, but not in 2. Hydrogen-deuterium (H/D) exchange rates determined by electrospray ionization mass spectrometry, however, decrease in the order 1 much greater than 2 > 3, while pH titrations reveal that the stability of the model protein folds decreases in the order 3 > 2 much greater than 1. Furthermore, H-1 NMR spectra of 2-Co and 3-Co indicate that the aromatic side chains in each compound are oriented within the shielding region of the porphyrin ring. Two-dimensional NOE and chemical shift data show that the helices in 3-Co are more highly organized than in 1-Co and span nearly the entire peptide sequence, while in 2-Co shorter helices of intermediate stability run between Phe-4 and Ala-13. The combined results indicate that aromatic side chain-porphyrin interactions in 2 and 3 stabilize their respective model protein folds, and suggest a similar role for the corresponding interactions in natural hemoproteins. Finally, the chemical shift patterns of the Trp side chains in 3-Co, the different effects of Phe and Trp on peptide architecture, and the pattern of chemical shifts exhibited by the alpha-NH and alpha-CH hydrogens in all three Co(III) PSMs demonstrate that the solution structures of these designed hemoproteins are similar to those predicted in molecular modeling studies.