从人骨肉瘤组织中纯化及鉴定一种病毒特异性逆转录酶的生化特性

Purification and biochemical characterization of a virus-specific reverse transcriptase from human osteosarcoma tissue.

作者信息

Ebener U, Welte K, Chandra P

出版信息

Cancer Lett. 1979 Aug;7(4):179-88. doi: 10.1016/s0304-3835(79)80078-6.

Abstract

A RNA-dependent DNA polymerase (RTase) was purified from human osteosarcoma tissue by successive column chromatography of the microsomal fraction on DEAE-cellulose (DE-23 and DE-52) and phosphocellulose. The purified enzyme has a molecular weight of about 68,000, a pH optimum of 8.1, a Mg2+ optimum of 0.8 mM, Mn2+ optimum of 1.0 mM and a KCl optimum of 60 mM. The enzyme transcribes (rA)n . (dT)12, (rC)n . (dG)12-18 and (2-O-methyl C)n . (dG)18, but is unable to transcribe (dA)n . (dT)10. The enzyme has no catalytic activity in the presence of oligodeoxynucleotide initiators alone, indicating the absence of terminal deoxynucleotidyl transferase. The purified enzyme is able to transcribe the heteropolymeric regions of a 70S RNA from R(Mu)LV. The presented data support the presence of a RNA-dependent DNA polymerase in human osteosarcoma tissue with biochemical properties, resembling those of C-type RNA tumor viruses.

摘要

通过将微粒体部分在DEAE - 纤维素（DE - 23和DE - 52）和磷酸纤维素上连续进行柱色谱，从人骨肉瘤组织中纯化出一种RNA依赖性DNA聚合酶（逆转录酶）。纯化后的酶分子量约为68,000，最适pH为8.1，最适Mg2 +浓度为0.8 mM，最适Mn2 +浓度为1.0 mM，最适KCl浓度为60 mM。该酶能转录（rA）n·（dT）12、（rC）n·（dG）12 - 18和（2 - O - 甲基C）n·（dG）18，但不能转录（dA）n·（dT）10。该酶在仅存在寡脱氧核苷酸引发剂时无催化活性，表明不存在末端脱氧核苷酸转移酶。纯化后的酶能够转录来自R（Mu）LV的70S RNA的杂聚区域。所呈现的数据支持人骨肉瘤组织中存在一种RNA依赖性DNA聚合酶，其生化特性与C型RNA肿瘤病毒的相似。