Basic Research for the Creation of Artificial Nucleic Acids and Related Biomolecules with Useful Functions

Hayakawa开发了高效固相合成DNA和RNA齐聚物的方法，该方法使用酸/唑络合物作为促进剂。他还发明了一种新的亚磷酰胺方法，使用分子筛作为除湿剂，允许使用化学计量的原料和促进剂在溶液中制备短寡核苷酸。这些新方法使得高效合成具有生物吸引力的重要物质成为可能，如cMP-Neu5Ac、2-5A core、c(GpGp)、脱氧核糖核酸-肽结合物以及以S-乙酰基-2-硫乙基三酯为骨架的寡核苷酸。Makino研究了未解决的核酸结构及其与生物功能的相关性。他(1)建立了一氧化氮对所有碱基的损伤及其与癌症和细胞死亡的关系的完整机制，(2)在人类端粒中发现了C-重复序列I-基序的各种异构体结构，并提示其与癌症和细胞死亡的关系，(3)阐明了硫代磷酸反义核酸的手性与其靶DNA和RNA识别能力的关系，(4)建立了针对给定碱基序列的肽库的新方法。Sekine发现，在H-膦酸法中，与寡核苷酸合成相关的严重副反应是由于生长链的5‘-末端羟基对最接近的核苷酸间磷酰基的分子内攻击。基于这一发现，一种使用活性酯中间体的改进的H-膦酸法已被开发出来。“质子封闭法”作为一种无碱基保护和后处理的DNA片段合成新策略也得到了探索。这一策略被用于获得N-酰化DNA片段的创新路线，这些片段已被证明具有足够的碱基配对能力。

Hayakawa has developed efficient solid-phase synthesis of DNA and RNA oligomers via the phosphoramidite method using an acid/azole complex as the promoter. He has also invented a novel phosphoramidite method with molecular sieves as a moisture scavenger, allowing the solution-phase preparation of short oligonucleotides by the use of stoichiometric amounts of starting materials and a promoter. These new approaches have allowed highly efficient synthesis of biologically attractive/important substances such as CMP-Neu5Ac, 2-5A core, c(GpGp), DNA-peptide conjugates, and oligodeoxyribonucleotides with S-acetyl-2-thioethyl triester backbone. Makino has studied to elucidate unsolved nucleic acids structures and its relevance to biological functions. He (1) has established complete mechanisms for all the base damage by nitric oxide and their relevance to cancer and cell death, (2) has found varied isomeric structures of i-motif of C-repeat in human telomere and suggested its relationship with cancer and cell death, (3) has elucidated the relationship between the chirality in phosphorothioate antisenses and their target DNA and RNA recognition ability, and (4) has developed the novel method for peptide library for given base sequences. Sekine has found that a serious side reaction associated with oligonucleotide synthesis in the H-phosphonate approach occurs owing to an intramolecular attack of the 5'-terminal hydroxyl group of the growing chain on the closest internucleotidic phosphoryl group. Based on this finding, an improved H-phosphonate approach using active ester intermediates has been developed. The "proton-block method" as a new strategy for the synthesis of DNA fragments without base protection and post-treatment has been also explored. This strategy was utilized for an innovative route to N-acylated DNA fragments which have proved to have sufficient base-pairing ability.