Integration of Non-natural Amino Acids Into Protein

1)设计合成了生物素基赖氨酰pCpA，使其整合到蛋白质中后更易于检测; 2）一种有效的制备含有任意核苷酸序列的反密码子且缺少3 '端pCpCpA的tRNA的方法，利用显微外科技术和酵母tRNA^<Phe>、tRNA^<Tyr>等技术，建立了一种非放射性测定低分子量RNA核苷酸序列的方法。3）含硫代磷酸的mRNA在核酸酶抗性和翻译效率方面的优势在大肠杆菌无细胞翻译系统中，证明了硫代mRNA相对于天然mRNA的表达。在极端嗜热菌Thermus themophilus HB 8的无细胞系统中，研究了不寻常的多胺对苯丙氨酰-tRNA形成的影响。还考察了酵母、圆柱假丝酵母和牛大肠杆菌的细胞树翻译系统的基本特征。

1) Several protecting groups of 5'-terminal and internucleotide phosphates and base amines were tested to obtain better yield of aminoacyl pCpCpA.Biotinyl-lysyl pCpA was designed and synthesized to make it easier to detect the amino acid after integration into proteins.2) An efficient method for preparing tRNA which has an anticodon of arbitrary nucleotide sequence and lacks 3'-terminal pCpCpA,was developed by the use of mictosurgery technique and yeast tRNA^<Phe>, tRNA^<Tyr> and so on. Furthermore, a non-radioactive method for determining nucleotide sequences of low-molecular-weight RNA was developed.3) Advantage in nuclease resistance and translational efficiency of phosphorothioate-containing mRNA (thio-mRNA) over natural mRNA was demonstrated in E.coli cell-free translation system. Effects of unusual polyamines on phenylalanyl-tRNA formation was investigated in cell-free systems of an extreme thermophile, Thermus themophilus HB8. Fundamental characteristics of cell-tree translation systems from yeast, Candida cylindracea and bovine mitochondriawere also examined.