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本文利用小白鼠脑或肝全细胞抽提液为酶源,Oligo A为底物,对真核哺乳动物细胞中的RNA连接酶进行研究.经测定,其抗碱性磷酸单醋酶的活性为16—49mU标准/ml.另外,还利用小白鼠脑或肝细胞核抽提液为酶源,人工合成的UpCpU和~(32)pNp为底物,用同系层析和放射自显影技术进行检测,得到清晰的连接产物自显影点.使用 DEAE-Sephadex A_(25)柱将连接产物进行了分离,并用同系层析法对连接产物做了进一步鉴定.用KOH或碱性磷酸单酯酶水解法,证明了连接方式为:3′-P→5′-OH.实验初步发现:在小白鼠脑或肝细胞核中有一与 T_4RNA连接酶不同的新酶。 相似文献
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RNA ligase in eukaryotic mammalian cells was studied by using mouse brain and livercell extracts as enzyme sources and Oligo A as substrates. RNA ligase activity was deter-mined by measuring the formation of alkaline phosphate-resistant product from 5'-~32P-termi-hated Oligoribonucleotides. Under appropriate conditions, the activity of this enzyme inbrain and liver cells may vary between 16-49 mU/ml. The joining way between donorand acceptor is 5'-P→3'-OH. Further studies wore carried out by using synthetic UpCpUand ~32pNp as substrates and crude enzyme preparations from extracts of cell unclei of brainand liver as enzyme sources. RNA ligase activity was examined by homochromatographyand autoradiography. A clear joining product was demonstrated and then isolated from thereaction mixture by DEAE-Sephadex A25 column chromatography. The eluted fractionswere identified by DEAE-cellulose thin layer chromatography. The joining product washydrolyzed either with KOH or with alkaline phosphatase, the autoradiog 相似文献
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