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首页> 外文期刊>The protein journal >Insufficient (sub-native) helix content in soluble/solid aggregates of recombinant and engineered forms of IL-2 throws light on how aggregated IL-2 is biologically active
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Insufficient (sub-native) helix content in soluble/solid aggregates of recombinant and engineered forms of IL-2 throws light on how aggregated IL-2 is biologically active

机译：重组和工程化形式的IL-2的可溶性/固体聚集体中螺旋含量不足（亚天然），这说明聚集的IL-2的生物活性如何

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Interleukin 2 (IL-2) is an extremely aggregation-prone, all-alpha helical cytokine. In its receptor-bound state, ～72 % of the polypeptide chain adopts helical structure and there is no beta sheet content whatsoever. In the past, recombinant IL-2 has been formulated and used therapeutically in humans, following production in E. coli. Therapeutic IL-2 consists entirely of functionally-active soluble aggregates with ～30 subunits per aggregate particle. Side-effects attributed to aggregation resulted in discontinuation of usage over a decade ago. Structurally, and biochemically, activity in IL-2 aggregates can potentially be explained in one of two ways: (a) individual IL-2 chains exist in sterically-accessible, receptor binding-competent (native) structures, allowing aggregates to bind directly to IL-2 receptors (IL-2R); alternatively, (b) IL-2 chains dissociate from aggregates, become free to adopt native structure, and then bind to IL-2R. We produced native IL-2 and numerous engineered forms in E. coli with the objective of obtaining insights into these possibilities. Each IL-2 variant was subjected to size exclusion chromatography, circular dichroism (CD) and Fourier transform infrared spectroscopy (FTIR). All forms produced and studied (including those with native IL-2 sequences) turned out to aggregate and also display less than ～50 % helix content as well as significant beta sheet content. No conditions were found that obviate aggregation. Aggregated IL-2 is thus insufficiently native-like to bind to IL-2R. Activity in aggregates thus probably owes to adoption of receptor binding-competent structures by chains that have already dissociated from aggregates.
机译：白介素2（IL-2）是一种极易聚集的全α螺旋细胞因子。在其受体结合状态下，约72％的多肽链采用螺旋结构，没有任何β折叠。过去，重组IL-2已在大肠杆菌中生产后配制并用于人体治疗。治疗性IL-2完全由具有功能活性的可溶性聚集体组成，每个聚集体颗粒含约30个亚基。归因于聚集的副作用导致十年前停止使用。从结构上和生物化学上，IL-2聚集体的活性可以用以下两种方式之一解释：（a）单个IL-2链以空间可及，受体结合能力（天然）结构存在，从而使聚集体直接结合至IL-2受体（IL-2R）;或者，（b）IL-2链从聚集物中解离，变得自由地采用天然结构，然后与IL-2R结合。我们在大肠杆菌中生产了天然IL-2和多种工程形式，目的是深入了解这些可能性。对每种IL-2变体进行尺寸排阻色谱，圆二色性（CD）和傅立叶变换红外光谱（FTIR）。产生和研究的所有形式（包括具有天然IL-2序列的形式）都聚集在一起，并且还显示出不到〜50％的螺旋含量以及显着的β折叠含量。没有发现消除聚集的条件。因此，聚集的IL-2天然不足以结合IL-2R。因此，聚集物中的活性可能是由于已经从聚集物中解离的链采用了具有受体结合能力的结构。

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