Crestoni Maria Elisa, Fornarini Simonetta
Dipartimento Studi di Chimica e Tecnologia delle Sostanze Biologicamente Attive, Università di Roma La Sapienza, P.le A. Moro 5, 00185, Rome, Italy.
J Biol Inorg Chem. 2007 Jan;12(1):22-35. doi: 10.1007/s00775-006-0159-9. Epub 2006 Aug 31.
High-resolution Fourier transform ion cyclotron resonance mass spectrometry is employed to gain thorough kinetics and thermodynamics information on the reaction of free and ligated heme-type ions with selected ligands, with the aim of obtaining an insight into the coordination environment of the prosthetic group in a variety of biomolecular ions. Adopting a stepwise approach towards systems of increasing complexity, we examined the reactivity of free gaseous iron(III) protoporphyrin IX ions, Fe(III)-heme(+), of the charged species from microperoxidase-11 (MP11) (covalently peptide bound heme), and of the multiply charged ions from heme proteins, namely, cytochrome c (cyt c) and myoglobin (examples of noncovalently protein bound hemes). Among an array of test compounds allowed to react with Fe(III)-heme(+), OP(OMe)(3) and P(OMe)(3) proved to be similarly efficient ligands in the first addition step, yet displayed markedly distinct reactivity towards heme iron already engaged in axial coordination. The ease with which P(OMe)(3) acts as a second axial ligand is exploited to probe structural and conformational features of biomolecular ions. In this way, circumstantial evidence is gained of a folded conformation of +2 charge state ions from MP11 and an elongated one for the +3 charge state ions. Similarly, both the general reaction pattern and detailed kinetics and thermodynamics data point to a regiospecific addition reaction of P(OMe)(3) directed at the heme iron within multiply charged ions from cyt c. This unprecedented example of ion-molecule reaction which specifically involves a prosthetic group belonging to protein ions stands in contrast to the multiple, nonspecific interactions established by OP(OMe)(3) molecules with the protonated sites of multiply charged cyt c and apomyoglobin ions. This finding may develop and provide sensitive probes of the structure and bonding features of protein ions in the gas phase.
采用高分辨率傅里叶变换离子回旋共振质谱法来获取关于游离和配位血红素型离子与选定配体反应的全面动力学和热力学信息，目的是深入了解各种生物分子离子中辅基的配位环境。我们采用逐步研究的方法，针对复杂度不断增加的体系，研究了游离气态铁（III）原卟啉IX离子（Fe(III)-血红素(+)）、微过氧化物酶-11（MP11）（共价肽结合血红素）的带电物种以及血红素蛋白的多电荷离子（即细胞色素c（cyt c）和肌红蛋白（非共价蛋白结合血红素的例子））的反应活性。在一系列与Fe(III)-血红素(+)反应的测试化合物中，OP(OMe)(3)和P(OMe)(3)在第一步加成反应中被证明是同样有效的配体，但对已经参与轴向配位的血红素铁显示出明显不同的反应活性。利用P(OMe)(3)作为第二个轴向配体的容易程度来探测生物分子离子的结构和构象特征。通过这种方式，获得了间接证据，证明MP11的 +2电荷态离子呈折叠构象，而 +3电荷态离子呈伸长构象。同样，一般反应模式以及详细的动力学和热力学数据都表明，P(OMe)(3)对cyt c的多电荷离子中的血红素铁进行区域特异性加成反应。这个前所未有的离子 - 分子反应例子特别涉及蛋白质离子中的辅基，这与OP(OMe)(3)分子与多电荷cyt c和脱辅基肌红蛋白离子的质子化位点建立的多种非特异性相互作用形成对比。这一发现可能会发展并提供气相中蛋白质离子结构和键合特征的灵敏探针。
