Structural analyses combined with small-angle X-ray scattering reveals that the retention of heme is critical for maintaining the structure of horseradish peroxidase under denaturing conditions
Structural analyses combined with small-angle X-ray scattering reveals that the retention of heme is critical for maintaining the structure of horseradish peroxidase under denaturing conditions
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DOI：
10.1007/s00726-016-2372-3
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发表时间：
2017-04-01
期刊：
影响因子：
3.5
通讯作者：
Choi, Kwan Yong
中科院分区：
文献类型：
--
作者：
Cha, Hyung Jin;Jang, Do Soo;Choi, Kwan Yong
我们分析了辣根过氧化物酶（HRP）在9 M尿素或6 M盐酸胍（GdnHCl）变性条件下的结构。远紫外圆二色性（CD）光谱表明存在天然的二级结构的holo-HRP在9 M尿素。此外，在近紫外和Soret区域CD光谱的holo-HRP在9 M尿素的轻微变化表明，holo-HRP的三级结构和血红素的结合在这种条件下保持部分完整。在9 M尿素中的holo-HRP的热展开过渡曲线中的过渡表明存在相当数量的二级结构。然而，没有二级结构，三级结构，或血红素和HRP之间的相互作用，观察在holo-HRP在6 M GdnHCl。小角X-射线散射表明，虽然在9 M尿素的holo-HRP的远端和近端域可能是部分展开，包含血红素的中心区域可能保持其三级结构。我们的研究结果表明，保留的血红素是必不可少的高度变性条件下的HRP的结构的维护。
We analyzed the structure of horseradish peroxidase (HRP) under denaturing conditions of 9 M urea or 6 M guanidine hydrochloride (GdnHCl). Far-UV circular dichroism (CD) spectra indicated the existence of native-like secondary structure of holo-HRP in 9 M urea. In addition, slight changes in near-UV and Soret region CD spectra of holo-HRP in 9 M urea suggest that the tertiary structure of holo-HRP and the binding of heme remain partially intact in this condition. A transition in the thermal unfolding transition curve of holo-HRP in 9 M urea indicated the existence of a considerable amount of secondary structure. However, no secondary structure, tertiary structure, or interaction between heme and HRP were observed in holo-HRP in 6 M GdnHCl. Small-angle X-ray scattering indicated that although distal and proximal domains of holo-HRP in 9 M urea might be partially unfolded, the central region that contains the heme might maintain its tertiary structure. Our results suggest that retention of the heme is essential for maintenance of the structure of HRP under highly denaturing conditions.