Suzuki H, Itano H A
Proc Natl Acad Sci U S A. 1973 Jul;70(7):2059-63. doi: 10.1073/pnas.70.7.2059.
Synthetic alphaT1 and betaT1, the N-terminal tryptic peptides of alpha-chain and beta-chain of hemoglobin, and MetalphaT1 and MetbetaT1, peptides in which N-terminal methionyl residues are peptide-bonded to alphaT1 and betaT1, were prepared by the solid-state method of Merrifield. These synthetic peptides were used to establish conditions for chromatographic purification and analysis. When tryptic digests of nascent globin chains from rabbit and sickle-cell anemia reticulocytes incubated with (35)S- and (3)H-labeled amino acids were analyzed, radioactivity not present in tryptic digests of labeled hemoglobin appeared at the elution positions of MetalphaT1 and MetbetaT1. The fraction of nascent chains with N-terminal methionine was higher in sickle-cell anemia reticulocytes than in rabbit reticulocytes. If rate of peptide-chain elongation in polysomes is uniform, nascent human chains must attain a greater length before removal of the initial methionyl residue. Length of nascent chain at time of removal was calculated from two independent sets of data, one obtained from [(35)S]methionine incorporation into MetalphaT1, MetbetaT1, alphaT5, and betaT5, and the other obtained from [(3)H]lysine and [(3)H]valine incorporation into MetbetaT1 and betaT1.
通过梅里菲尔德的固相法制备了合成的αT1和βT1（血红蛋白α链和β链的N端胰蛋白酶肽段）以及MetαT1和MetβT1（其中N端甲硫氨酰残基通过肽键与αT1和βT1相连的肽段）。这些合成肽用于建立色谱纯化和分析的条件。当分析用（35）S和（3）H标记的氨基酸孵育的兔和镰状细胞贫血网织红细胞新生珠蛋白链的胰蛋白酶消化产物时，标记血红蛋白的胰蛋白酶消化产物中不存在的放射性出现在MetαT1和MetβT1的洗脱位置。镰状细胞贫血网织红细胞中具有N端甲硫氨酸的新生链比例高于兔网织红细胞。如果多核糖体中肽链延伸速率是均匀的，那么新生的人链在去除初始甲硫氨酰残基之前必须达到更长的长度。去除时新生链的长度是根据两组独立数据计算得出的，一组数据来自将[（35）S]甲硫氨酸掺入MetαT1、MetβT1、αT5和βT5，另一组数据来自将[（3）H]赖氨酸和[（3）H]缬氨酸掺入MetβT1和βT1。
