A RESTRICTION ENZYME FROM HEMOPHILUS-INFLUENZAE .2. BASE SEQUENCE OF RECOGNITION SITE
A RESTRICTION ENZYME FROM HEMOPHILUS-INFLUENZAE .2. BASE SEQUENCE OF RECOGNITION SITE
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DOI：
10.1016/0022-2836(70)90150-6
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发表时间：
1970-01-01
影响因子：
5.6
通讯作者：
SMITH, HO
中科院分区：
文献类型：
--
作者：
KELLY, TJ;SMITH, HO
流感嗜血杆菌菌株Rd含有一种酶，内切核酸酶R，它特异性地降解外源DNA。以噬菌体T7 DNA为底物，内切核酸酶引入有限数量（约40个）双链断裂（5′-磷酰基，3′-羟基）。极限产物的平均长度约为1000个核苷酸对，不含单链断裂。我们通过标记5′-磷酰基（使用多核苷酸激酶）并表征由各种核酸酶释放的标记片段，探索了极限产物5′-末端的核苷酸序列。得到两类5′-末端序列：pApApCpNp.（60%）和pGpApCpNp.（40%），其中N表示第4位碱基不是唯一的。对限制性产物进行微球菌核酸酶消化后，分离出3′端的二核苷单磷酸，并鉴定为TpT（60%）和TpC（40%）。我们得出结论，核酸内切酶R的H。流感病毒识别的特异性核苷酸序列为：5′... pGpTpPy β pPupApCp.3 ′... pCpApPup β PypTpGp.5 ′。
Hemophilus influenzaestrain Rd contains an enzyme, endonuolease R, which specifically degrades foreign DNA. With phage T7 DNA as substrate the endonuclease introduces a limited number (about 40) double-strand breaks (5′-phosphoryl, 3′-hydroxyl). The limit product has an average length of about 1000 nucleotide pairs and contains no single-strand breaks. We have explored the nucleotide sequences at the 5′-ends of the limit product by labeling the 5′- phosphoryl groups (using polynucleotide kinase) and characterizing the labeled fragments released by various nucleases. Two classes of 5′-terminal sequences were obtained: pApApCpNp … (60%) and pGpApCpNp … (40%), where N indicates that the base in the 4th position is not unique. The dinucleoside monophosphates at the 3′-ends were isolated after micrococcal nuclease digestion of the limit product and identified as TpT(60%) and TpC (40%). We conclude that endonuclease R ofH. influenzaerecognizes the following specific nucleotide sequence: 5′ … pGpTpPy ¦pPupApCp … 3′ 3′ … pCpApPup ¦PypTpGp … 5′ The implications of the twofold rotational symmetry of this sequence are discussed.