Enzymatic synthesis of <sup>3</sup>H-thymidine-5-triphosphate

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Enzymatic synthesis of ³H-thymidine-5-triphosphate

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- Enzymatic synthesis of ³H-thymidine-5-triphosphate
- Nuclear Techniques Vol. 05, Issue 1, (1982)
- 作者机构：
  
  中国科学院上海细胞生物学研究所
- 作者简介：
- 基金信息：
- DOI：
  CLC：
- Received：04 August 1980，
  
  Published：1982-02
- 稿件说明：
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Enzymatic synthesis of ³H-thymidine-5-triphosphate[J]. Nuclear techniques, 1982, 05(1): 37-40,91.
DOI：

Enzymatic synthesis of ³H-thymidine-5-triphosphate[J]. Nuclear techniques, 1982, 05(1): 37-40,91. DOI：

摘要

在生命科学的一些基础研究方面，如DNA的复制机制，mRNA的反向转录和遗传工程等，

H-甲基-胸腺嘧啶核苷三磷酸（

H-TPP）是一种不可少的标记化合物。

H-TTP的酶促合成方法是以

H-TdR作为底物，在ATP和Mg

存在的条件下，先后通过TdR激酶、TMP和TDP的激酶作用、酶促磷酸化来合成

H-TTP

[1]

。上述各种激酶，可以从不同的来源得到，如Smellie等人

[2]

是从艾氏腹水细胞中提取；Baugnet-Mahieu等人

[3]

是用大白鼠再生肝来提取。Bessman等人

[4]

则从大肠肝菌中获得。我们采用前两种方法并与酵母中的酶系

[7，8]

进行比较，摸索了

H-TTP酶促合成的最适条件，获得了转化率达75%以上、纯度高、分离简便的合成方法。

Abstract

It was shown that the crude extracts of Ehrlich ascites carcinoma which contain the activities of thymidine kinase and thymidine monophosphate kinase and diphosphate kinase can effectively catalyze the synthetic reaction of

H-TTP from the substracte—

H-thymidine (TdR). The conversion rate of TdR to TTP by these extracts was more than 75%

higher than those by rat regenerating liver and yeast. A method for complete separation of

H-TdR and

H-TTP was established using DEAE-Sephadex A-25 column chromatography

and then eluting with ammonium bicarbonate. The product was salt free. The optimum conditions for the reaction were ATP and enzyme Mg

(11µmol each)

reaction time 2hr. at 37℃ and enzyme protein

less than 5mg per milliliter reaction volume.

关键词

Keywords

references

S. M. Weissman et al . , Biochimica et Biophysica Acta , 45 , 101 ( 1960 ).

R. M. S. Smcllie et al . , Biochimica et Biophysica Acta , 35 , 389 ( 1959 ).

L. Baugnet-Mahieu et al . , J. Labelled Compounds , 2 , 77 ( 1966 ).

Maurice J. Bessman et al . , J. Biol. Chem . , 233 , 171 ( 1958 ).

H. M. Keir and R. M. S. Smellie , Biochimica et Biophysica , 35 , 405 ( 1959 ).

E. D. Gray et al . , Biochimica et Biophysica Acta , 45 , 111 ( 1960 ).

胡兆庆等，微生物学报， 18 , 3 , 239 （ 1978 ）.

中国科学院微生物研究所病毒变别组和上海原子核研究所氚标记组，生物化学与生物物理学报， 9 , 2 , 201 （ 1977 ）.

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⁰

Enzymatic synthesis of 3H-thymidine-5-triphosphate

DOI：

摘要

Abstract

关键词

Keywords

references

Enzymatic synthesis of ³H-thymidine-5-triphosphate