The attachment of a DNA-binding Sso7d-like protein improves processivity and resistance to inhibitors of M-MuLV reverse transcriptase

Igor P. Oscorbin, Pei Fong Wong, Ulyana A. Boyarskikh, Evgeny A. Khrapov, Maksim L. Filipenko

Результат исследования: Научные публикации в периодических изданияхстатьярецензирование

Аннотация

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.

Язык оригиналаанглийский
Число страниц19
ЖурналFEBS Letters
Ранняя дата в режиме онлайн24 сен 2020
DOI
СостояниеЭлектронная публикация перед печатью - 24 сен 2020

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