Yarbrough D, Wachter R M, Kallio K, Matz M V, Remington S J
Departments of Physics and Biology and the Institute of Molecular Biology, University of Oregon, Eugene, OR 97403, USA
Proc Natl Acad Sci U S A. 2001 Jan 16;98(2):462-7. doi: 10.1073/pnas.98.2.462.
The crystal structure of DsRed, a red fluorescent protein from a corallimorpharian, has been determined at 2.0-A resolution by multiple-wavelength anomalous dispersion and crystallographic refinement. Crystals of the selenomethionine-substituted protein have space group P2(1) and contain a tetramer with 222 noncrystallographic symmetry in the asymmetric unit. The refined model has satisfactory stereochemistry and a final crystallographic R factor of 0.162. The protein, which forms an obligatory tetramer in solution and in the crystal, is a squat rectangular prism comprising four protomers whose fold is extremely similar to that of the Aequorea victoria green fluorescent protein despite low ( approximately 23%) amino acid sequence homology. The monomer consists of an 11-stranded beta barrel with a coaxial helix. The chromophores, formed from the primary sequence -Gln-Tyr-Gly- (residues 66-68), are arranged in a approximately 27 x 34-A rectangular array in two approximately antiparallel pairs. The geometry at the alpha carbon of Gln-66 (refined without stereochemical restraints) is consistent with an sp(2) hybridized center, in accord with the proposal that red fluorescence is because of an additional oxidation step that forms an acylimine extension to the chromophore [Gross, L. A., Baird, G. S., Hoffman, R. C., Baldridge, K. K. & Tsien, R. Y. (2000) Proc. Natl. Acad. Sci. USA 87, 11990-11995]. The carbonyl oxygen of Phe-65 is almost 90 degrees out of the plane of the chromophore, consistent with theoretical calculations suggesting that this is the minimum energy conformation of this moiety despite the conjugation of this group with the rest of the chromophore.
通过多波长反常色散和晶体学精修，已在2.0埃分辨率下测定了来自珊瑚虫红荧光蛋白DsRed的晶体结构。硒代蛋氨酸取代蛋白的晶体属于空间群P2(1)，在不对称单元中含有一个具有222非晶体学对称性的四聚体。精修后的模型具有令人满意的立体化学结构，最终晶体学R因子为0.162。该蛋白在溶液和晶体中均形成必需的四聚体，是一个矮胖的长方体棱柱，由四个原体组成，尽管氨基酸序列同源性较低（约23%），但其折叠方式与维多利亚多管水母绿色荧光蛋白极为相似。单体由一个带有同轴螺旋的11股β桶组成。发色团由一级序列-Gln-Tyr-Gly-（残基66 - 68）形成，以两个近似反平行的对排列在一个约27×34埃的矩形阵列中。Gln-66的α碳处的几何结构（在无立体化学限制下精修）与sp(2)杂化中心一致，这与红色荧光是由于发色团形成酰亚胺延伸的额外氧化步骤的提议相符[格罗斯，L.A.，贝尔德，G.S.，霍夫曼，R.C.，鲍德里奇，K.K.和钱永健，R.Y.（2000年）美国国家科学院院刊87，11990 - 11995]。Phe-65的羰基氧几乎与发色团平面成90度角，这与理论计算结果一致，表明尽管该基团与发色团的其余部分共轭，但这是该部分的最低能量构象。
