Folkers P J, van Duynhoven J P, Jonker A J, Harmsen B J, Konings R N, Hilbers C W
Nijmegen SON Research Center, University of Nijmegen, The Netherlands.
Eur J Biochem. 1991 Dec 5;202(2):349-60. doi: 10.1111/j.1432-1033.1991.tb16382.x.
Sequence-specific 1H-NMR assignments are reported for the Tyr41----His (Y41H) mutant of the single-stranded DNA binding protein, encoded by gene V of the filamentous bacteriophage M13 (GVP). The mutant protein was chosen for this purpose because it exhibits significantly improved solubility characteristics over wild-type GVP [Folkers et al. (1991) Eur. J. Biochem. 200, 139-148]. The secondary structure elements present in the protein are deduced from a qualitative interpretation of the nuclear Overhauser enhancement spectra and amide exchange data. The protein is entirely composed of antiparallel beta-structure. It is shown that identical structural elements are present in wild-type GVP. Previously, we have demonstrated that the secondary structure of the beta-loop, encompassing residues 13-31 which is present in GVP in solution, deviates from that proposed for the same amino acid sequence on the basis of X-ray diffraction data [van Duynhoven et al. (1990) FEBS Lett. 261, 1-4]. Now that we have arrived at a complete description of the secondary structure of the protein in solution, other deviations with respect to the crystallographically determined structure became apparent as well. The N-terminal part of the protein is, in solution, part of a triple-stranded beta-sheet while, in the crystal, it is an extended strand pointing away from the bulk of the protein dimer. One of the antiparallel beta-sheets in the protein which had been designated earlier as the complex loop has, in the solution structure, a different pairwise arrangement of the residues in its respective beta-ladders. Residues 30 and 48 are opposite to one another in the solution structure while in the crystal structure residues 32 and 48 are paired. A similar observation is made for the so-called dyad domain of the protein of which the beta-sheet in the solution structure is shifted by one residue with respect to that of the crystal structure.
报道了丝状噬菌体M13基因V编码的单链DNA结合蛋白的Tyr41→His（Y41H）突变体的序列特异性1H-NMR归属。选择该突变蛋白进行此项研究是因为它比野生型GVP具有显著改善的溶解性[Folkers等人（1991年），《欧洲生物化学杂志》200，139 - 148]。蛋白质中存在的二级结构元件是通过对核Overhauser增强光谱和酰胺交换数据的定性解释推导出来的。该蛋白质完全由反平行β结构组成。结果表明野生型GVP中存在相同的结构元件。此前，我们已经证明，溶液中GVP中包含13 - 31位残基的β环的二级结构与基于X射线衍射数据对相同氨基酸序列所提出的结构不同[van Duynhoven等人（1990年），《欧洲生物化学学会联合会快报》261，1 - 4]。既然我们已经得到了该蛋白质在溶液中的二级结构的完整描述，那么与晶体学确定的结构相比的其他差异也变得明显了。在溶液中，该蛋白质的N端部分是三链β折叠的一部分，而在晶体中，它是一条远离蛋白质二聚体主体的延伸链。该蛋白质中一个较早被指定为复合环的反平行β折叠在溶液结构中其各自β梯中的残基具有不同的配对排列。在溶液结构中30位和48位残基彼此相对，而在晶体结构中32位和48位残基配对。对于该蛋白质的所谓二联体结构域也有类似的观察结果，其溶液结构中的β折叠相对于晶体结构的β折叠偏移了一个残基。
