Search: (hsv:(NATURVETENSKAP) hsv:(Biologi) hsv:(Biokemi och molekylärbiologi)) pers:(Uhlen Mathias)
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Production of a The...
Production of a Thermostable DNA Polymerase by Site-Specific Cleavage of a Heat-Eluted Affinity Fusion Protein
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- Gräslund, Torbjörn (author)
- KTH,Biokemi och biokemisk teknologi
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- Nilsson, Joakim (author)
- KTH,Biokemi och biokemisk teknologi
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Lindberg, Michael (author)
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- Uhlén, Mathias (author)
- KTH,Biokemi och biokemisk teknologi
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- Nygren, Per-Åke (author)
- KTH,Biokemi och biokemisk teknologi
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(creator_code:org_t)
- Elsevier BV, 1997
- 1997
- English.
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In: Protein Expression and Purification. - : Elsevier BV. - 1046-5928 .- 1096-0279. ; 9, s. 125-132
- Related links:
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http://www.sciencedi...
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https://urn.kb.se/re...
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https://doi.org/10.1...
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Abstract
Subject headings
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- A novel strategy is described for bacterial expression and affinity purification of a recombinant truncated version of the heat-stable DNA polymerase I fromThermus aquaticus.The DNA polymerase ([Delta]Taq) was produced as a fusion to a serum albumin binding affinity handle (ABP) derived from streptococcal protein G. Based on the thermostability of the [Delta]TaqDNA polymerase, affinity-purified ABP-[Delta]Taqcould be heat-eluted from HSA columns by incubation at 85ï¿œC. To produce free [Delta]TaqDNA polymerase, efficient site-specific cleavage of the affinity tag was performed using a recombinant coxsackievirus 3C protease (3Cpro), also produced as an ABP affinity fusion. Thus, an integrated strategy could be devised where both the cleaved ABP affinity tag and the protease fusion could be recovered after site-specific cleavage using HSA-affinity chromatography. The flow-through fraction contained essentially pure [Delta]TaqDNA polymerase with full enzymatic activity.
Subject headings
- NATURVETENSKAP -- Biologi -- Biokemi och molekylärbiologi (hsv//swe)
- NATURAL SCIENCES -- Biological Sciences -- Biochemistry and Molecular Biology (hsv//eng)
Keyword
- Biochemistry
- Biokemi
Publication and Content Type
- ref (subject category)
- art (subject category)
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