一种用甲醛和硫醇对腺嘌呤核苷酸中N6-氨基进行化学修饰的新方法及其在制备固定化ADP和ATP中的应用。

A new method of chemical modification of N6-amino group in adenine nucleotides with formaldehyde and a thiol and its application to preparing immobilized ADP and ATP.

作者信息

Yamazaki Y, Suzuki H

出版信息

Eur J Biochem. 1978 Dec 1;92(1):197-207. doi: 10.1111/j.1432-1033.1978.tb12738.x.

DOI:10.1111/j.1432-1033.1978.tb12738.x

Abstract

Reaction of AMP with formaldehyde and 3-mercaptopropionic acid at pH 11.7 gave a new AMP derivative, N6-[(2-carboxyethyl)thiomethyl]-AMP (I) in 91% yield and reaction at pH 3.1 gave another new derivative, N6,N6-bis[(2-carboxyethyl)thiomethyl]-AMP (II) in 57% yield. The structures were determined by their 13C and 1H nuclear magnetic resonance spectra coupled with those of the simple analogues, N6-[(2-carboxyethyl)thiomethyl]-9-methyladenine (III) and N6,N6-bis[(2-carboxyethyl)thiomethyl]-9-methyladenine (IV) which were synthesized from 9-methyladenine in the same way as for derivatives I and II. ADP and ATP were treated in the same way as AMP to afford the corresponding carboxyl derivatives, N6-[(2-carboxyethyl)thiomethyl]-ADP (V), N6-[(2-carboxyethyl)thiomethyl]-ATP (VI), N6,N6-bis[(2-carboxyethyl)thiomethyl]-ADP (X) and N6,N6-bis[(2-carboxyethyl)thiomethyl]-ATP (XI) in 71%, 75%, 53% and 40% yield, respectively. These compounds were coupled to 1,3-diaminopropane with a water-soluble carbodiimide to give the corresponding amino derivatives, N6-([N-3-aminopropyl)carbamoylethyl]thiomethyl)-ADP (VIII), N6-(N-(3-aminopropyl)carbamoylethyl]thiomethyl)-ATP (IX), N6,N6-bis([N-(3-aminopropyl)carbamoylethyl]thiomethyl)-ADP (XIII), and N6,N6-bis([N-(3-aminopropyl)carbamoylethyl]thiomethyl)-ATP (XIV), which were further bound to CNBr-activated dextran to give new polymer-bound derivatives of ADP and ATP. These free and bo-nd derivatives were tested for their coenzymic activities against several kinases. The activities of the ADP derivatives, V, VIII, X, XIII, dextran-bound VIII, and dextran-bound XIII against acetate kinase were 82%, 81%, 68%, 55%, 35%, and 15%, respectively, relative to ADP and those of the ATP derivatives, VI, IX, XI, XIV, dextran-bound IX, and dextran-bound XIV against hexokinase were 88%, 94%, 60%, 81%, 58%, and 49%, respectively, relative to ATP.

摘要

在pH 11.7条件下，AMP与甲醛和3-巯基丙酸反应生成一种新的AMP衍生物，N6-[(2-羧乙基)硫代甲基]-AMP（I），产率为91%；在pH 3.1条件下反应生成另一种新衍生物，N6,N6-双[(2-羧乙基)硫代甲基]-AMP（II），产率为57%。通过13C和1H核磁共振光谱以及简单类似物N6-[(2-羧乙基)硫代甲基]-9-甲基腺嘌呤（III）和N6,N6-双[(2-羧乙基)硫代甲基]-9-甲基腺嘌呤（IV）的光谱确定了其结构，这两种类似物是由9-甲基腺嘌呤以与衍生物I和II相同的方式合成的。ADP和ATP以与AMP相同的方式处理，分别得到相应的羧基衍生物，N6-[(2-羧乙基)硫代甲基]-ADP（V）、N6-[(2-羧乙基)硫代甲基]-ATP（VI）、N6,N6-双[(2-羧乙基)硫代甲基]-ADP（X）和N6,N6-双[(2-羧乙基)硫代甲基]-ATP（XI），产率分别为71%、75%、53%和40%。这些化合物与1,3-二氨基丙烷用一种水溶性碳二亚胺偶联，得到相应的氨基衍生物，N6-([N-3-氨基丙基)氨甲酰乙基]硫代甲基)-ADP（VIII）、N6-(N-(3-氨基丙基)氨甲酰乙基]硫代甲基)-ATP（IX）、N6,N6-双([N-(3-氨基丙基)氨甲酰乙基]硫代甲基)-ADP（XIII）和N6,N6-双([N-(3-氨基丙基)氨甲酰乙基]硫代甲基)-ATP（XIV），它们进一步与CNBr活化的葡聚糖结合，得到ADP和ATP的新的聚合物结合衍生物。测试了这些游离和结合衍生物对几种激酶的辅酶活性。相对于ADP，ADP衍生物V、VIII、X、XIII、葡聚糖结合的VIII和葡聚糖结合的XIII对乙酸激酶的活性分别为82%、81%、68%、55%、35%和15%；相对于ATP，ATP衍生物VI、IX、XI、XIV、葡聚糖结合的IX和葡聚糖结合的XIV对己糖激酶的活性分别为88%、94%、60%、81%、58%和49%。