Title:
HELIX-HAIRPIN-HELIX MOTIFS TO MANIPULATE PROPERTIES OF DNA PROCESSING ENZYMES
Document Type and Number:
WIPO Patent Application WO2004013279
Kind Code:
A3
Abstract:
The utility of Topo V's HhH motifs for modulating DNA binding properties of a DNA processing enzyme, such as Taq DNA polymerase or PfuDNA polymerase, is demonstrated. In one embodiment, an amino acid residue comprising one or more HhH domains derived from Topoisomerase V, linked to either the NH2-terminus or COOH-terminus of a Taq polymerase fragment, significantly broadens the processivity and/or salt concentration range of polymerase activity. The specific activities of the chimeric polymerases are not affected by added HhH motifs. Depending on the type of the construct, the thermal stability of chimeric polymerases increases or remains the same as that of Taq DNA polymerase or its Stoffel fragment. This invention further provides a method of increasing the salt tolerance of Taq polymerases. The methods of this invention may be applied to all pol A-type DNA polymerases, as well as to other DNA processing enzymes.
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Inventors:
SLESAREV ALEXEI (US)
PAVLOV ANDREY (US)
KOZYAVKIN SERGEI (US)
PAVLOVA NADEZHDA (US)
PAVLOV ANDREY (US)
KOZYAVKIN SERGEI (US)
PAVLOVA NADEZHDA (US)
Application Number:
PCT/US2003/015336
Publication Date:
October 07, 2004
Filing Date:
May 14, 2003
Export Citation:
Assignee:
FIDELITY SYSTEMS INC (US)
SLESAREV ALEXEI (US)
PAVLOV ANDREY (US)
KOZYAVKIN SERGEI (US)
PAVLOVA NADEZHDA (US)
SLESAREV ALEXEI (US)
PAVLOV ANDREY (US)
KOZYAVKIN SERGEI (US)
PAVLOVA NADEZHDA (US)
International Classes:
C07K14/195; A61K39/00; A61K48/00; (IPC1-7): C12Q1/68; C12P19/34
Domestic Patent References:
| WO2001092501A1 | 2001-12-06 | |||
| WO1999047649A2 | 1999-09-23 |
Other References:
NIXON A.E. ET AL: "Hybrid enzymes: manipulating enzyme design", TIBTECH, vol. 16, no. 6, June 1998 (1998-06-01), pages 258 - 264, XP004121067
BELOVA G.I. ET AL: "The domain organization and properties of individual domains of DNA topoisomerase V, a type 1B topoisomerase with DNA repair activities", J. BIOL. CHEM., vol. 277, no. 7, December 2001 (2001-12-01), pages 4959 - 4965, XP002978491
BELOVA G.I. ET AL: "The domain organization and properties of individual domains of DNA topoisomerase V, a type 1B topoisomerase with DNA repair activities", J. BIOL. CHEM., vol. 277, no. 7, December 2001 (2001-12-01), pages 4959 - 4965, XP002978491
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