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首页> 外文期刊>Biochemistry >Circular permutation of granulocyte colony-stimulating factor.
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Circular permutation of granulocyte colony-stimulating factor.

机译：粒细胞集落刺激因子的圆形排列。

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The sequence of granulocyte colony-stimulating factor (G-CSF) has been circularly permuted by introducing new chain termini into interhelical loops and by constraining the N- and C-terminal helices, either by direct linkage of the termini (L0) or by substitution of the amino-terminal 10-residue segment with a seven-residue linker composed of glycines and serines (L1). All the circularly permuted G-CSFs (cpG-CSFs) were able to fold into biologically active structures that could recognize the G-CSF receptor. CD and NMR spectroscopy demonstrated that all of the cpG-CSFs adopted a fold similar to that of the native molecule, except for one [cpG-CSF(L1)[142/141]] which has the new termini at the end of loop 34 with the shorter L1 linker. All of the cpG-CSFs underwent cooperative unfolding by urea, and a systematically lower free energy change (DeltaGurea) was observed for molecules with the shorter L1 linker than for those molecules in which the original termini were directly linked (the L0 linker). The thermodynamic stability of the cpG-CSFs toward urea was found to correlate with their relative ability to stimulate proliferation of G-CSF responsive cells. Taken together, these results indicate that the G-CSF sequence is robust in its ability to undergo linear rearrangement and adopt a biologically active conformation. The choice of linker, with its effect on stability, seems to be important for realizing the full biological activity of the three-dimensional structure. The breakpoint and linker together are the ultimate determinants of the structural and biological profiles of these circularly permuted cytokines. In the following paper [McWherter, C. A., et al. (1999) Biochemistry 38, 4564-4571], McWherter and co-workers have used circularly permuted G-CSF sequences to engineer chimeric dual IL-3 and G-CSF receptor agonists in which the relative spatial orientation of the receptor agonist domains is varied. Interpreting the differences in activity for the chimeric molecules in terms of the connectivity between domains depends critically on the results reported here for the isolated cpG-CSF domains.
机译：通过将新的链末端引入螺旋间环并通过末端的直接连接（L0）或通过取代来限制N和C末端螺旋，已对粒细胞集落刺激因子（G-CSF）的序列进行了循环排列氨基末端的10个残基片段具有一个由甘氨酸和丝氨酸组成的7个残基的连接子（L1）。所有环状排列的G-CSF（cpG-CSF）都可以折叠成可以识别G-CSF受体的生物活性结构。 CD和NMR谱图表明，除了一个[cpG-CSF（L1）[142/141]]在环34末端具有新末端的[cpG-CSF（L1）[142/141]]之外，所有cpG-CSF都采用了与天然分子相似的折叠方式。使用较短的L1接头。所有cpG-CSF都通过尿素进行协同解折叠，观察到具有L1接头较短的分子比那些直接连接了原始末端的分子（L0接头）具有更低的自由能变化（DeltaGurea）。发现cpG-CSF对尿素的热力学稳定性与其刺激G-CSF反应性细胞增殖的相对能力相关。综上所述，这些结果表明G-CSF序列在进行线性重排和采用生物活性构象方面具有强大的能力。连接子的选择及其对稳定性的影响，似乎对于实现三维结构的全部生物活性很重要。断点和接头一起是这些环状排列的细胞因子的结构和生物学特征的最终决定因素。在以下论文中[McWherter，C. A.，et al。 （1999）Biochemistry 38，4564-4571]，McWherter和同事使用了环状排列的G-CSF序列来设计嵌合双IL-3和G-CSF受体激动剂，其中受体激动剂结构域的相对空间方向发生了变化。根据域之间的连通性来解释嵌合分子的活性差异主要取决于此处报道的针对分离的cpG-CSF域的结果。

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