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当前位置: 首页 > 文章 > 长角血蜱唾液腺中腺苷三磷酸双磷酸酶的纯化及其酶切机制 昆虫学报 2002,45 (3) 307-312
Position: Home > Articles > Purification and mechanism of enzymic hydrolysis of apyrase from salivary glands of Haemaphysalis longicornis Acta Entomologica Sinica 2002,45 (3) 307-312

长角血蜱唾液腺中腺苷三磷酸双磷酸酶的纯化及其酶切机制

作  者：
程远国;吴厚永;李德昌;李成文;赵彤言
单  位：
军事医学科学院微生物与流行病研究所;解放军军需大学寄生虫学教研室
关键词：
长角血蜱;唾液腺;腺苷三磷酸双磷酸酶;酶水解
摘  要：
利用染料亲和层析 (CibacornBlue柱 )和离子交换层析 (MacrosphereWCX柱 )对长角血蜱Haemaphysalislongicornis唾液腺的腺苷三磷酸双磷酸酶进行纯化 ,经SDS PAGE证实其分子量为 6 6kD。腺苷三磷酸双磷酸酶可以水解ATP和ADP ,但对AMP无水解作用 ,水解ATP和ADP的Km 值均为 0 2 μmol L ,Vmax值分别为 12 5和 15 6 μmol (min·mg)。腺苷三磷酸双磷酸酶水解ATP的中间产物是ADP ,最终产物是AMP和正磷酸。表明腺苷三磷酸双磷酸酶水解ATP的位点是 5′ 核苷酸的γ 磷酸键 ,水解ADP的位点是 5′ 核苷酸的 β 磷酸键。
译  名：
Purification and mechanism of enzymic hydrolysis of apyrase from salivary glands of Haemaphysalis longicornis
作  者：
CHENG Yuan-Guo 1, WU Hou-Yong\+1,LI De-Chang\+2, LI Cheng-Wen\+1, ZHAO Tong-Yan\+1 (1. Institute of Microbiology and Epidemiology, Beijing\ 100071, China; 2. Military Necessaries University, Changchun\ 130042, China)
关键词：
Haemaphysalis longicornis; salivary gland; apyrase; enzymic hydrolysis
摘  要：
Apyrase was isolated from soluble crude salivary glands extract of \%Haemaphysalis longicornis\% by a Cibacron Blue column and a Macrosphere weak cation-exchange column. Apyrase could hydrolyse ATP and ADP, but failed to hydrolyse AMP. The approach K m, obtained with ATP and ADP as substrates, were evaluated 0.2 μmol/L and the approach V max were 12.5 and 15.6 μmol/(min·mg) respectively. During the hydrolysis of ATP as monitored by HPLC, the UV-absorbing compounds observed were the ADP and AMP,and for ADP was the AMP only. Direct evidence showed that the hydrolysis site of ATP and ADP occurred by cleavage at the phosphate residues γ and β to the 5′ carbon atom of the ribose was obtained by enzymically hydrolyzing both [γ- 32 P] ATP and [β- 32 P] ADP and separating the products by ion-exchange HPLC. The mechanism of ATP and ADP hydrolysis is sequential removal of phosphate residues to produce, finally, AMP.

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