PROBING THE STABILITY OF A PARTLY FOLDED APOMYOGLOBIN INTERMEDIATE BY SITE-DIRECTED MUTAGENESIS
PROBING THE STABILITY OF A PARTLY FOLDED APOMYOGLOBIN INTERMEDIATE BY SITE-DIRECTED MUTAGENESIS
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DOI：
10.1021/bi00231a001
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发表时间：
1991-04-30
期刊：
影响因子：
2.9
通讯作者：
BALDWIN, RL
中科院分区：
文献类型：
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作者：
HUGHSON, FM;BARRICK, D;BALDWIN, RL
部分折叠形式（I）的脱辅基肌红蛋白的α-螺旋含量约为35%;在早期的研究中，氢交换揭示了A、G和H螺旋是折叠的，而蛋白质的大部分其余部分不是[休森，F. M.，赖特，体育，& Baldwin，R. L.（1990）Science 249，1544-1548]。 由于A，G和H形成一个紧凑的子域在本地肌红蛋白，我们提出，nativelike包装之间的相互作用的三个螺旋可能会保留在I型的脱辅基肌红蛋白。 为了测试这个提议，破坏性突变被引入到A. H和G. H螺旋包装位点。 这些突变使天然脱辅基肌红蛋白相对于I不稳定。 相反，I的稳定性对突变相对不敏感;特别是，单独的侧链体积似乎并不重要。 这些结果表明，I型并不能通过天然的AH和GH包装相互作用来稳定。 在支持这一点，我们表明，部分螺旋肽来自肌红蛋白的G和H螺旋区不配对在溶液中。 由于分离的G肽和H肽最多只是部分螺旋，因此某种类型的相互作用必须使这些螺旋稳定在I型。 当突变引入非极性表面积增加的侧链时，可以看到I的稳定性略有增加。 我们认为，我是稳定的相对非特异性的疏水相互作用，使其能够很容易地适应突变。 在这方面和其他方面，I似乎符合“熔融球”模型，但需要说明的是，只有部分多肽链似乎参与了球。
A partly folded form (I) of apomyoglobin has an alpha-helix content of about 35%; in an earlier study, hydrogen exchange revealed that the A, G, and H helices are folded, while much of the rest of the protein is not [Hughson, F. M., Wright, P. E., & Baldwin, R. L. (1990) Science 249, 1544-1548]. Because A, G, and H form a compact subdomain in native myoglobin, we proposed that nativelike packing interactions among the three helices might be retained in the I form of apomyoglobin. To test this proposal, disruptive mutations were introduced into the A.H and G.H helix packing sites. These mutations destabilize native apomyoglobin relative to I. In contrast, the stability of I is relatively insensitive to mutation; in particular, side-chain volume alone does not appear to be important. These results indicate that the I form is not stabilized by nativelike A.H and G.H packing interactions. In support of this we show that partly helical peptides derived from the G and H helix regions of myoglobin do not pair in solution. Since the isolated G and H peptides are at best only partly helical, some type of interaction must stabilize these helices in the I form. Small increases in the stability of I are seen when mutation introduces a side chain of increased nonpolar surface area. We suggest that I is stabilized by relatively nonspecific hydrophobic interactions that allow it to adapt easily to mutation. In this and other respects, I appears to conform to the "molten globule" model, with the caveat that only part ofthe polypeptide chain appears to participate in the globule.