PEGylation Promotes Hemoglobin Tetramer Dissociation
PEGylation Promotes Hemoglobin Tetramer Dissociation
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DOI：
10.1021/bc900130f
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发表时间：
2009-07-01
影响因子：
4.7
通讯作者：
Mozzarelli, Andrea
中科院分区：
文献类型：
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作者：
Caccia, Dario;Ronda, Luca;Mozzarelli, Andrea
与聚（乙二醇）（PEG）缀合的血红蛋白充当血浆中游离的氧载体，在缺氧病理中替代红细胞补充氧。考虑到氧气输送控制的复杂性，PEG化血红蛋白的细微结构和功能差异可能与不同的生理反应和潜在的不良反应相关。我们比较了在厌氧条件下聚乙二醇化的血红蛋白（称为PEG-Hb（脱氧））与在有氧条件下聚乙二醇化的血红蛋白（称为PEG-Hb（氧合），一种模拟Hemospan的产品，由Sangart，Inc.生产）。SDS PAGE和MALDI-TOF分析表明，PEG缀合产生的产物的特征在于PEG/血红蛋白比率的广泛分布。尺寸排阻色谱中的洗脱曲线表明，在脱氧条件下和较高浓度下，两种产品均表现出更均匀的分子量/流体动力学体积分布。PEG-Hb（oxy）显示高氧亲和力、低变构效应物调节、几乎无协同作用、快速和双相CO结合。以及功能特性对浓度的有限依赖性，而PEG-Hb（脱氧）表现出显著依赖于蛋白质浓度的氧结合曲线，以及与天然血红蛋白相似的缓慢CO结合。聚乙二醇化CO-血红蛋白，探测闪光光解，表现出较低的振幅相对于原生血红蛋白和T状态双分子CO再结合相的双生再结合相。这些发现是。解释为聚乙二醇化血红蛋白解离成二聚体和扰动的T和R状态与降低的四元跃迁速率增加。这些特征对于PEG-Hb（oxy）比对于PEG-Hb（deoxy）更明显。当聚乙二醇化血红蛋白用作血液替代品时，检测到的异质性可能是不良反应的来源。
Hemoglobin conjugated with poly(ethylene glycol) (PEG) acts as an oxygen carrier free in plasma, substituting red blood cells in supplementing oxygen in hypo-oxygenation pathologies. Given the complexity of oxygen delivery controls, subtle structural and functional differences in PEGylated hemoglobins might be associated with distinct physiological responses and, potentially, adverse effects. We have compared hemoglobin PEGylated under anaerobic conditions, called PEG-Hb(deoxy), with hemoglobin PEGylated under aerobic conditions, called PEG-Hb(oxy), a product that mimics Hemospan, produced by Sangart, Inc. SDS PAGE and MALDI-TOF analyses demonstrated that PEG conjugation yields products characterized by a broad distribution of PEG/hemoglobin ratios. The elution profiles in size-exclusion chromatography indicate that both products exhibit a more homogeneous distribution of molecular weight/hydrodynamic volume under deoxy conditions and at higher concentrations. PEG-Hb(oxy) shows high oxygen affinity, low modulation of allosteric effectors, almost no cooperadvity, a fast and monophasic CO binding. and a limited dependence of functional properties on concentration, whereas PEG-Hb(deoxy) exhibits oxygen binding curves that significantly depend on protein concentration, and a slow CO binding, similar to native hemoglobin. PEGylated CO-hemoglobins, probed by flash photolysis, exhibited a lower amplitude for the geminate rebinding phase with respect to native hemoglobin and a negligible T state bimolecular CO rebinding phase. These findings are. explained by an increased dissociation of PEGylated hemoglobins into dimers and perturbed T and R states with decreased quaternary transition rates. These features are more pronounced for PEG-Hb(oxy) that) PEG-Hb(deoxy). The detected heterogeneity might be a source of adverse effects when PEGylated Hbs are used as blood substitutes.