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Charge engineering ...
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Gräslund, TorbjörnKTH,Biokemi och biokemisk teknologi
(författare)
Charge engineering of a protein domain to allow efficient ion-exchange recovery
- Artikel/kapitelEngelska2000
Förlag, utgivningsår, omfång ...
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2000-10
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Oxford University Press (OUP),2000
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printrdacarrier
Nummerbeteckningar
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LIBRIS-ID:oai:DiVA.org:kth-13275
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https://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-13275URI
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https://doi.org/10.1093/protein/13.10.703DOI
Kompletterande språkuppgifter
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Språk:engelska
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Sammanfattning på:engelska
Ingår i deldatabas
Klassifikation
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Ämneskategori:ref swepub-contenttype
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Ämneskategori:art swepub-publicationtype
Anmärkningar
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QC 20100609
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We have created protein domains with extreme surface charge. These mutated domains allow for ion-exchange chromatography under conditions favourable for selective and efficient capture, using Escherichia coli as a host organism. The staphylococcal protein A-derived domain Z (Z(wt)) was used asa scaffold when constructing two mutants, Z(basic1) and Z(basic2), with high positive surface charge. Far-ultraviolet circular dichroism measurements showed that they have a secondary structure content comparable to the parental molecule Z(wt). Although melting temperatures (T-m) of the engineered domains were lower than that of the wild-type Z domain, both mutants could be produced successfully as intracellular full-length products in E. coli and purified to homogeneity by ion-exchange chromatography. Further studies performed on Z(basic1) and Z(basic2) showed that they were able to bind to a cation exchanger even at pH values in the 9 to 11 range. A gene fusion between Z(basic2) and the acidic human serum albumin binding domain (ABD), derived from streptococcal protein G, was also constructed. The gene product Z(basic2)-ABD could be purified using cation-exchange chromatography from a whole cell lysate to more than 90% purity.
Ämnesord och genrebeteckningar
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NATURVETENSKAP Biologi hsv//swe
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NATURAL SCIENCES Biological Sciences hsv//eng
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circular dichroism
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ion-exchange chromatography
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molecular modelling
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pI
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protein A
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bacterial receptor domain
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escherichia-coli k-12
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fusion protein
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binding domain
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nucleic-acids
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force-field
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purification
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dna
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resolution
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sequence
Biuppslag (personer, institutioner, konferenser, titlar ...)
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Lundin, Gunnel
(författare)
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Uhlén, MathiasKTH,Biokemi och biokemisk teknologi(Swepub:kth)u1dulvmw
(författare)
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Nygren, Per-ÅkeKTH,Biokemi och biokemisk teknologi(Swepub:kth)u1zhverl
(författare)
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Hober, SophiaKTH,Biokemi och biokemisk teknologi(Swepub:kth)u11qqzc1
(författare)
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KTHBiokemi och biokemisk teknologi
(creator_code:org_t)
Sammanhörande titlar
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Ingår i:Protein Engineering: Oxford University Press (OUP)13:10, s. 703-7090269-21391460-213X
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Ingår i:Protein Engineering, Design and Selection: Oxford University Press (OUP)13:10, s. 703-7091741-01341741-0126
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