The attachment of a DNA‐binding Sso7d‐like protein improves processivity and resistance to inhibitors of M‐MuLV reverse transcriptase,FEBS Letters

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The attachment of a DNA‐binding Sso7d‐like protein improves processivity and resistance to inhibitors of M‐MuLV reverse transcriptase
FEBS Letters ( IF 3.0 ) Pub Date : 2020-10-05 , DOI: 10.1002/1873-3468.13934
Igor P. Oscorbin _{1,

2} , Pei Fong Wong _{1,

2} , Ulyana A. Boyarskikh _{1,

2} , Evgeny A. Khrapov ₁ , Maksim L. Filipenko _{1,

2}

Affiliation

Reverse transcriptases (RTs) are a standard tool in both fundamental studies and diagnostics. RTs should possess elevated temperature optimum, high thermal stability, processivity and tolerance to contaminants. Here, we constructed a set of chimeric RTs, based on the combination of the Moloney murine leukaemia virus (M‐MuLV) RT and either of two DNA‐binding domains: the DNA‐binding domain of the DNA ligase from Pyrococcus abyssi or the DNA‐binding Sto7d protein from Sulfolobus tokodaii. The processivity and efficiency of cDNA synthesis of the chimeric RT with Sto7d at the C‐end are increased several fold. The attachment of Sto7d enhances the tolerance of M‐MuLV RT to the most common amplification inhibitors: NaCl, urea, guanidinium chloride, formamide, components of human whole blood and human blood plasma. Thus, fusing M‐MuLV RT with an additional domain results in more robust and efficient RTs.

中文翻译：

DNA 结合 Sso7d 样蛋白的附着提高了持续合成能力和对 M-MuLV 逆转录酶抑制剂的抵抗力

逆转录酶 (RT) 是基础研究和诊断中的标准工具。RT 应具有最佳高温、高热稳定性、加工能力和对污染物的耐受性。在这里，我们构建了一组嵌合 RT，基于莫洛尼鼠白血病病毒 (M-MuLV) RT 和两个 DNA 结合域之一的组合：来自 Pyrococcus abyssi 的 DNA 连接酶的 DNA 结合域或 DNA结合来自 Sulfolobus tokodaii 的 Sto7d 蛋白。在 C 端具有 Sto7d 的嵌合 RT 的 cDNA 合成的持续合成能力和效率提高了几倍。Sto7d 的附着增强了 M-MuLV RT 对最常见扩增抑制剂的耐受性：NaCl、尿素、氯化胍、甲酰胺、人全血和人血浆成分。因此，

更新日期：2020-10-05

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