Hausinger R P, Howard J B
Proc Natl Acad Sci U S A. 1980 Jul;77(7):3826-30. doi: 10.1073/pnas.77.7.3826.
The molecular weights, amino acid compositions, amino- and carboxyl-terminal sequences, and ion-exchange peptide maps of the cysteine-containing tryptic peptides were determined for the iron proteins from the nitrogen fixation complexes of Azotobacter vinelandii (Av2) and Klebsiella pneumoniae (Kp2). Our results are compared to the known amino acid sequence of the iron protein from Clostridium pasteurianum (Cp2) [Tanaka, M., Haniu, M., Yasunobu, K. & Mortenson, L. E. (1977) J. Biol. Chem. 252, 7093-7100]. Previous studies have shown the iron proteins to have similar enzymatic functions and spectroscopic properties. Furthermore, the DNAs coding for the iron protein from many different species cross-hybridize [Ruvkun, G. B. & Ausubel, F. M. (1980) Proc. Natl. Acad. Sci. USA 77, 191-195]. Our results indicate that the protein structures are similar yet have significant differences. The amino-terminal sequences of Av2 and Kp2 are extended compared to the amino-terminal methionine of Cp2 and may indicate a different initiation site in these proteins. The aminoterminal sequences for Av2 and Kp2 are more homologous with each other than either of these are with Cp2. The carboxyl-terminal sequences are extended in Av2(14 residues) and Kp2 ( approximately 30 residues) compared to Cp2. The amino- and carboxyl-terminal sequences establish that either the structural gene sizes are different in the three organisms or extensive posttranslational modification must occur in some species. Because cysteinyl residues are involved at the active site of the iron protein, a sensitive peptide mapping technique was used to compare cysteinyl peptides of the iron protein from the three species. Av2 and Kp2 have a redistribution of cysteinyl residues when compared to Cp2. Three important differences in the cysteine distributions were found, namely, residue 4 is valine and residue 148 is alanine in Cp2, but cysteinyl residues occupy these positions in Av2, whereas residue 231 is cysteine in Cp2 but alanine in Av2. The peptide mapping technique provides a method for the investigation of selective chemical modification of cysteinyl residues.
测定了来自棕色固氮菌（Av2）和肺炎克雷伯氏菌（Kp2）固氮复合物中铁蛋白的分子量、氨基酸组成、氨基和羧基末端序列以及含半胱氨酸的胰蛋白酶肽的离子交换肽图谱。我们的结果与巴氏梭菌（Cp2）铁蛋白的已知氨基酸序列进行了比较[田中，M.，羽生，M.，安信，K.和莫滕森，L.E.（1977年）《生物化学杂志》252，7093 - 7100]。先前的研究表明铁蛋白具有相似的酶功能和光谱性质。此外，编码来自许多不同物种铁蛋白的DNA会发生交叉杂交[鲁夫昆，G.B.和奥苏贝尔，F.M.（1980年）《美国国家科学院院刊》77，191 - 195]。我们的结果表明蛋白质结构相似但存在显著差异。与Cp2的氨基末端甲硫氨酸相比，Av2和Kp2的氨基末端序列有所延长，这可能表明这些蛋白质中的起始位点不同。Av2和Kp2的氨基末端序列彼此之间比它们与Cp2的序列更同源。与Cp2相比，Av2（14个残基）和Kp2（约30个残基）的羧基末端序列有所延长。氨基和羧基末端序列表明，要么这三种生物体中的结构基因大小不同，要么在某些物种中必须发生广泛的翻译后修饰。由于半胱氨酰残基参与铁蛋白的活性位点，因此使用了一种灵敏的肽图谱技术来比较这三种物种铁蛋白的半胱氨酰肽。与Cp2相比，Av2和Kp2的半胱氨酰残基有重新分布。发现了半胱氨酸分布的三个重要差异，即Cp2中的第4位残基是缬氨酸，第148位残基是丙氨酸，但Av2中这些位置被半胱氨酰残基占据，而Cp2中的第231位残基是半胱氨酸，Av2中是丙氨酸。肽图谱技术为研究半胱氨酰残基的选择性化学修饰提供了一种方法。
