Polyols induce ATP-independent folding of GroEL-bound bacterial glutamine synthetase.
Polyols induce ATP-independent folding of GroEL-bound bacterial glutamine synthetase.
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DOI：
10.1006/abbi.2001.2620
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发表时间：
2002-01
影响因子：
3.9
通讯作者：
P. Voziyan;M. Fisher
中科院分区：
文献类型：
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作者：
P. Voziyan;M. Fisher
我们之前评估了细菌谷氨酰胺合成酶 (GS) 折叠的 GroE 伴侣蛋白要求，并确定，在 37°C 的 50 mM Tris 缓冲液中，ATP 与 GroEL-GS 复合物的结合对于十二聚酶的释放和重新激活是必需的。然而，我们在这里证明，在没有核苷酸的情况下，向 GroEL-GS 复合物中添加 1-4 M 甘油会导致 GS 的释放和重新激活。此外，这种甘油诱导的重折叠的重折叠动力学和重折叠产量与用 ATP 观察到的相似。其他多元醇如蔗糖、1,2-丙二醇或1,3-丙二醇也促进伴侣蛋白复合物中GS的不依赖于核苷酸的重折叠。观察到的现象不能归因于粘度或分子拥挤效应，因为与 4 M 甘油具有相同粘度的葡聚糖或聚蔗糖溶液未能重新激活 GroEL 结合的 GS。与甘油一样，其他渗透剂如甜菜碱和肌氨酸或高盐（500 mM NaCl）促进 GS 的自发折叠。然而，使用这些添加剂没有观察到结合 GroEL 的 GS 重新激活。甘油的存在影响荧光探针 1,8-苯胺萘与 GroEL 的结合，表明甘油可能改变伴侣蛋白结构。我们的数据表明，低分子量多元醇会影响 GroEL 和结合的 GS 单体，从而降低它们的结合亲和力。这导致 GS 向活跃的、有组装能力的状态的分配增加。
We have previously assessed the GroE chaperonin requirements for folding of bacterial glutamine synthetase (GS) and established that, at 37 degrees C in 50 mM Tris buffer, ATP binding to the GroEL-GS complex is mandatory for the release and reactivation of dodecameric enzyme. However, we demonstrate here that the addition of 1-4 M glycerol to GroEL-GS complexes resulted in release and reactivation of GS in the absence of nucleotide. Furthermore, the kinetics of refolding and refolding yields of this glycerol-induced refolding were similar to those observed with ATP. Other polyols such as sucrose, 1,2-propanediol, or 1,3-propanediol also facilitated nucleotide-independent refolding of GS from chaperonin complex. The observed phenomenon cannot be attributed to the viscosity or molecular crowding effects because solutions of dextran or Ficoll with the same viscosity as 4 M glycerol failed to reactivate GroEL-bound GS. Like glycerol, other osmolytes such as betaine and sarcosine or high salt (500 mM NaCl) facilitated spontaneous folding of GS. However, no reactivation of GroEL-bound GS was observed with these additives. The presence of glycerol affected binding of fluorescent probe 1,8-anilinonaphthalene to GroEL, suggesting that glycerol may alter the chaperonin structure. Our data suggest that low-molecular-weight polyols affect both GroEL and bound GS monomers to reduce their binding affinity. This results in an increased partitioning of GS toward active, assembly-competent states.