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

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The attachment of a DNA-binding Sso7d-like protein improves processivity and resistance to inhibitors of M-MuLV reverse transcriptase. / Oscorbin, Igor P.; Wong, Pei Fong; Boyarskikh, Ulyana A. et al.

In: FEBS Letters, Vol. 594, No. 24, 12.2020, p. 4338-4356.

Research output: Contribution to journal › Article › peer-review

BibTeX

@article{d9b4d6c05ab444928d04311863675b6a,

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

abstract = "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.",

keywords = "cDNA synthesis, chimeric protein, M-MuLV reverse transcriptase, protein engineering, reverse transcription, VIRUS, RNA, PERFORMANCE, POLYMERASE, AVIAN-MYELOBLASTOSIS, AMPLIFICATION, INCREASE, GENERATION, THERMOSTABILITY, RIBONUCLEASE",

author = "Oscorbin, {Igor P.} and Wong, {Pei Fong} and Boyarskikh, {Ulyana A.} and Khrapov, {Evgeny A.} and Filipenko, {Maksim L.}",

note = "The research was supported by the Russian State Funded Budget Project of ICBFM. Funding for the open access charge: Russian Science Foundation.",

year = "2020",

month = dec,

doi = "10.1002/1873-3468.13934",

language = "English",

volume = "594",

pages = "4338--4356",

journal = "FEBS Letters",

issn = "0014-5793",

publisher = "Wiley-Blackwell",

number = "24",

}

RIS

TY - JOUR

T1 - The attachment of a DNA-binding Sso7d-like protein improves processivity and resistance to inhibitors of M-MuLV reverse transcriptase

AU - Oscorbin, Igor P.

AU - Wong, Pei Fong

AU - Boyarskikh, Ulyana A.

AU - Khrapov, Evgeny A.

AU - Filipenko, Maksim L.

N1 - The research was supported by the Russian State Funded Budget Project of ICBFM. Funding for the open access charge: Russian Science Foundation.

PY - 2020/12

Y1 - 2020/12

N2 - 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.

AB - 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.

KW - cDNA synthesis

KW - chimeric protein

KW - M-MuLV reverse transcriptase

KW - protein engineering

KW - reverse transcription

KW - VIRUS

KW - RNA

KW - PERFORMANCE

KW - POLYMERASE

KW - AVIAN-MYELOBLASTOSIS

KW - AMPLIFICATION

KW - INCREASE

KW - GENERATION

KW - THERMOSTABILITY

KW - RIBONUCLEASE

UR - http://www.scopus.com/inward/record.url?scp=85092027910&partnerID=8YFLogxK

U2 - 10.1002/1873-3468.13934

DO - 10.1002/1873-3468.13934

M3 - Article

C2 - 32970841

AN - SCOPUS:85092027910

VL - 594

SP - 4338

EP - 4356

JO - FEBS Letters

JF - FEBS Letters

SN - 0014-5793

IS - 24

ER -

ID: 25677196