Hunt R E, Pratt L H
Biochemistry. 1980 Jan 22;19(2):390-4. doi: 10.1021/bi00543a022.
We characterized immunoaffinity-purified, undegraded oat (Avena sativa L., cv. Garry) phytochrome by several physicochemical techniques. Phytochrome, of greater than 98% purity [Hunt, R. E., & Pratt, L. H. (1979) Plant Physiol. 64, 332-336], existed in solution as a dimer of its 118 000-dalton monomers and had a full complement of the typical amino acids with about 35% nonpolar residues, 115 carboxylic acid groups per monomer, and an average of one phosphate per monomer. Although the dimer was not held together by disulfide bridges, each monomer contained three disulfide bonds and 14 reduced cysteines out of a total of 27 cysteine-half-cystine residues. Phytochrome preparations, although very pure, exhibited heterogeneity by discontinuous sodium dodecyl sulfate-polyacrylamide gel electrophoresis, which revealed three closely spaced bands, and by nondenaturing gel electrophoresis at pH 7.0, which revealed four bands. Amino-terminal analysis indicated two residues, Lys and Ala. Manual Edman degradation yielded Leu and Ala after one round and Val and Leu after a second round. These data indicate a possible amino-terminal sequence of NH3-Lys-Ala-Leu-Val- with some monomers not having Lys.
我们运用多种物理化学技术对免疫亲和纯化、未降解的燕麦（燕麦属，栽培品种Garry）光敏色素进行了表征。纯度大于98%的光敏色素[亨特，R.E.，&普拉特，L.H.（1979年）《植物生理学》64卷，332 - 336页]，在溶液中以其118000道尔顿单体的二聚体形式存在，具有完整的典型氨基酸组成，约35%为非极性残基，每个单体有115个羧酸基团，且每个单体平均有一个磷酸基团。尽管二聚体并非通过二硫键维系在一起，但每个单体含有三个二硫键以及总共27个半胱氨酸 - 半胱氨酸残基中的14个还原型半胱氨酸。光敏色素制剂尽管纯度很高，但通过不连续十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳显示出异质性，该电泳揭示了三条紧密相邻的条带，而在pH 7.0的非变性凝胶电泳中则揭示了四条条带。氨基末端分析表明有两个残基，赖氨酸和丙氨酸。手动埃德曼降解在第一轮后得到亮氨酸和丙氨酸，第二轮后得到缬氨酸和亮氨酸。这些数据表明可能的氨基末端序列为NH3 - 赖氨酸 - 丙氨酸 - 亮氨酸 - 缬氨酸 - ，有些单体不含赖氨酸。
