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首页> 外文期刊>Acta crystallographica, Section F. Structural biology and crystallization communications >Crystallographic structure determination of B10 mutants of Vitreoscilla hemoglobin: role of Tyr29 (B10) in the structure of the ligand-binding site
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Crystallographic structure determination of B10 mutants of Vitreoscilla hemoglobin: role of Tyr29 (B10) in the structure of the ligand-binding site

机译：玻璃体血红蛋白 B10 突变体的晶体结构测定：Tyr29 （B10） 在配体结合位点结构中的作用

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摘要

Site-directed mutants of the gene encoding wild-type Vitreoscilla hemoglobin were made that changed Tyr29 (B10) of the wild-type Vitreoscilla hemoglobin (VHb) to either Phe or Ala. The wild-type and the two mutant hemoglobins were expressed in Escherichia coli and purified to homogeneity. The binding of the two mutants to CO was essentially identical to that of wild-type VHb as determined by CO-difference spectra. Circular-dichroism spectra also showed the two mutants to be essentially the same as wild-type VHb regarding overall helicity. All three VHbs were crystallized and their structures were determined at resolutions of 1.71.9 angstrom, which are similar to that of the original wild-type structure determination. The Tyr29Phe mutant has a structure that is essentially indistinguishable from that of the wild type. However, the structure of the Tyr29Ala mutant has significant differences from that of the wild type. In addition, for the Tyr29Ala mutant it was possible to determine the positions of most of the residues in the D region, which was disordered in the originally reported structure of wild-type VHb as well as in the wild-type VHb structure reported here. In the Tyr29Ala mutant, the five-membered ring of proline E8 (Pro54) occupies the space occupied by the aromatic ring of Tyr29 in the wild-type structure. These results are discussed in the context of the proposed role of Tyr29 in the structure of the oxygen-binding pocket.
机译：编码野生型玻璃体血红蛋白的基因的定点突变体被制成，将野生型玻璃体血红蛋白 （VHb） 的 Tyr29 （B10） 改变为 Phe 或 Ala。野生型血红蛋白和两种突变血红蛋白在大肠杆菌中表达并纯化至均一性。两种突变体与CO的结合与野生型VHb的结合基本相同，由CO差谱确定。圆二色谱图也显示，这两种突变体在整体螺旋度方面与野生型VHb基本相同。3种VHb均结晶，其结构分辨率为1.71.9埃，与原野生型结构测定结果相似。Tyr29Phe突变体的结构与野生型基本没有区别。然而，Tyr29Ala突变体的结构与野生型有显著差异。此外，对于Tyr29Ala突变体，可以确定D区中大多数残基的位置，该残基在最初报道的野生型VHb结构以及本文报道的野生型VHb结构中是无序的。在Tyr29Ala突变体中，脯氨酸E8（Pro54）的五元环占据了野生型结构中Tyr29芳香环所占据的空间。这些结果在Tyr29在氧结合口袋结构中的作用的背景下进行了讨论。

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