Biochemical characterization of Epstein-Barr virus nuclear antigen 2A and an associated ATPase activity.
Biochemical characterization of Epstein-Barr virus nuclear antigen 2A and an associated ATPase activity.
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Epstein-Barr 病毒核抗原 2A 的生化特征和相关的 ATP 酶活性。
DOI：
10.1111/j.1432-1033.1992.tb17019.x
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发表时间：
1992
期刊：
影响因子：
--
通讯作者：
Klein,G
中科院分区：
文献类型：
--
作者：
Randahl,H;Fåhraeus,R;Klein,G
A partial purification of the Epstein‐Barr‐virus nuclear antigen 2A (EBNA 2A) protein from the Epstein‐Barr‐virus‐infected lymphoblastoid cell line, Cherry, has been designed. The main purification step was immunoaffinity chromatography, based on the mAb, 115E, directed towards the carboxy terminus of EBNA 2A. This was followed by chromatography over a Blue Sepharose column. According to silver‐stained SDS/PAGE, EBNA 2A was estimated to be 20% pure. The purified fractions contained an ATPase activity that was inhibited by the mAb 115E. Immunopurification of six EBNA‐2A‐positive cell lines and their negative counterpart showed that only fractions from EBNA‐2A‐positive lines contained ATPase activity. In gel‐filtration experiments EBNA 2A eluted as a 75‐kDa protein in conjunction with an ATPase activity. The EBNA 2A protein was covalently labeled by the ATP analog [14C]5′‐[p‐(fluorosulfonyl)benzoyl]adenosine. The ATPase activity was found to be optimal in the presence of 0.25 mM MgCl2or CaCl2, whereas, in the presence of MnCl2and ZnCl2, the activity was only about 50% of the control. High concentrations of Na2VO3and heparin do not interfere with the activity, while 2.5 mM NaF or 0.5 M NaCl give a 50% reduction of the activity. TheKmfor ATP and for GTP was 13 μM and 11 μM, respectively, and theVmaxfor ATP was about six‐times higher than with GTP as substrate. Other low‐molecular‐mass non‐protein phosphate esters, such as phosphoserine or phosphothreonine inhibited the ATPase activity with aKiof 18 and 32 μM, respectively. Phosphotyrosine had aKiof 480 μM. Serine, threonine and tyrosine had no inhibitory effect on the ATPase activity.
