The effect of a point mutation on the stability of IgG4 as monitored by analytical ultracentrifugation
The effect of a point mutation on the stability of IgG4 as monitored by analytical ultracentrifugation
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DOI：
10.1002/jps.21016
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发表时间：
2008-02-01
影响因子：
3.8
通讯作者：
Mulot, Sandrine
中科院分区：
文献类型：
--
作者：
Lu, Yanling;Harding, Stephen E.;Mulot, Sandrine
目前人们对抗体制剂的聚集状态和生物物理完整性有相当大的兴趣，最近在分析性超速离心数据分析方面的进展使其成为在储存和冻融条件下这些稳定性现象的强有力的探索者。将野生型IgG4抗体和单一氨基酸铰链突变体IgG4m(丝氨酸241位转变为脯氨酸)的溶液暴露在不同的加速应力条件下，即(I)不同时期的高温储存(37℃下长达59天)或(Ii)一系列冻融循环(在-80℃下储存，然后在不同条件下在20℃下孵育1h)。在分析超速离心机中使用无干扰的沉降速度探针进行的分析表明，无论使用何种储存方式，对于这两种抗体而言，单体始终是最常见的物种。沉降系数分布分析表明，存在其他较高的寡聚体种类和半抗体，它们之间似乎不处于化学平衡状态。溶液的异质性被发现在处理天然抗体和铰链突变抗体时显著增加，尽管后者似乎对冻融诱导的聚集更具抵抗力。(C)2007年Wiley-Liss，Inc.
There is presently considerable interest in the state of aggregation and biophysical integrity of antibody preparations, and recent advances in the analysis of data from the analytical ultracentrifuge renders it a powerful probe of these stability phenomena, under both storage and freeze-thaw conditions. Solutions of a wildtype IgG4 antibody and a single amino acid hinge mutant IgG4m (serine residue 241 converted to proline) were exposed to different accelerated stress conditions, namely (i) elevated temperature storage for various periods (up to 59 days at 37 degrees C) or (ii) a series of freeze-thaw cycles (storage at - 80 degrees C then incubation at 20 degrees C for 1 h under different conditions). Analysis using the nondisruptive probe of sedimentation velocity in the analytical ultracentrifuge indicated that for both antibodies the monomer was always the most common species present whatever storage regime had been used. Sedimentation coefficient distribution analysis showed that other higher oligomer species and half-antibodies were present, and appeared to be not in chemical equilibrium with each other. Solution heterogeneity was found to increase considerably with treatment for both native and hinge-mutant antibodies although the latter appeared to be more resistant to freeze-thaw-induced aggregation. (c) 2007 Wiley-Liss, Inc.