A carbohydrate binding module as a diversity-carrying scaffoldShow others and affiliations
2004 (English)In: Protein Engineering Design & Selection, ISSN 1741-0126, E-ISSN 1741-0134, Vol. 17, no 3, p. 213-221Article in journal (Refereed) Published
Abstract [en]
The growing field of biotechnology is in constant need of binding proteins with novel properties. Not just binding specificities and affinities but also structural stability and productivity are important characteristics for the purpose of large-scale applications. In order to find such molecules, libraries are created by diversifying naturally occurring binding proteins, which in those cases serve as scaffolds. In this study, we investigated the use of a thermostable carbohydrate binding module, CBM4-2, from a xylanase found in Rhodothermus marinus, as a diversity-carrying scaffold. A combinatorial library was created by introducing restricted variation at 12 positions in the carbohydrate binding site of the CBM4-2. Despite the small size of the library (1.6 x 10(6) clones), variants specific towards different carbohydrate polymers (birchwood xylan, Avicel and ivory nut mannan) as well as a glycoprotein (human IgG4) were successfully selected for, using the phage display method. Investigated clones showed a high productivity (on average 69 mg of purified protein/l shake flask culture) when produced in Escherichia coli and they were all stable molecules displaying a high melting transition temperature (75.7 +/- 5.3 degrees C). All our results demonstrate that the CBM4-2 molecule is a suitable scaffold for creating variants useful in different biotechnological applications.
Place, publisher, year, edition, pages
Oxford University Press, 2004. Vol. 17, no 3, p. 213-221
Keywords [en]
binding specificity; carbohydrate binding module; molecular library; phage selection; scaffold
National Category
Biochemistry Molecular Biology
Identifiers
URN: urn:nbn:se:oru:diva-64487DOI: 10.1093/protein/gzh026ISI: 000222797300003PubMedID: 15082834Scopus ID: 2-s2.0-2942748131OAI: oai:DiVA.org:oru-64487DiVA, id: diva2:1177099
2018-01-242018-01-242025-02-20Bibliographically approved