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- Hedin, Eva. M. K.KTH,Bioteknologi (author)
Selective reduction and chemical modification of oxidized lipase cysteine mutants
- Article/chapterEnglish2002
Publisher, publication year, extent ...
- Canadian Science Publishing,2002
- printrdacarrier
Numbers
- LIBRIS-ID:oai:DiVA.org:kth-21725
- https://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-21725URI
- https://doi.org/10.1139/v02-046DOI
Supplementary language notes
- Language:English
- Summary in:English
Part of subdatabase
- SwePubSwePub
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Notes
- QC 20100525
- Thirteen single-cysteine mutants of the 33 kDa fungal triacylglycerol lipase Thermomyces (formerly Humicola) lanuginosa lipase (TLL, EC 3.1.1.3) Were produced and characterized for the purpose of site-directed chemical modification with spectroscopic reporter groups. All cysteine mutants were found to be predominantly blocked by oxidation to disulfides with endogenous cysteine during production. The fraction of lipase molecules with free sulfhydryl groups was analyzed by labeling with N-biotinylaminoethyl methanethiosulfonate, followed by a novel dot-blot method based on biotin-streptavidin interactions. A non-invasive method for the reduction of the introduced cysteine was elaborated for this protein containing three native disulfide bridges. The site-specifically reduced TLL mutants were then labeled with the sulfhydryl-specific reagents 2-(5-dimethylaminonaphth-1-ylsulfonamido)ethyl methanethiosulfonate or (1-oxyl-2,2,5,5-tetramethyl-Delta(3)-pyrroline-3-methyl) methanethiosulfonate, and studied by fluorescence and electron spin resonance (ESR) spectroscopy.
Subject headings and genre
- MEDICIN OCH HÄLSOVETENSKAP Medicinsk bioteknologi Medicinsk bioteknologi hsv//swe
- MEDICAL AND HEALTH SCIENCES Medical Biotechnology Medical Biotechnology hsv//eng
- lipase
- cysteine mutant
- selective reduction
- chemical modification
- methanethiosulfonate
- humicola-lanuginosa lipase
- fluorescence spectroscopy
- conformational-changes
- protein
- stability
- sulfhydryl
- dynamics
- enzymes
- binding
- thiols
- Biochemistry
- Biokemi
Added entries (persons, corporate bodies, meetings, titles ...)
- Patkar, S. A.KTH,Bioteknologi (author)
- Vind, J.KTH,Bioteknologi (author)
- Svendsen, A.KTH,Bioteknologi (author)
- Hult, KarlKTH,Biokemi och biokemisk teknologi(Swepub:kth)u1nzlwuj (author)
- Berglund, PerKTH,Biokemi och biokemisk teknologi(Swepub:kth)u1271chh (author)
- KTHBioteknologi (creator_code:org_t)
Related titles
- In:Canadian journal of chemistry (Print): Canadian Science Publishing80:6, s. 529-5390008-40421480-3291
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