Biophysical characterization of Z(SPA-1)--a phage-display selected binder to protein A

Protein Science : a Publication of the Protein Society
Christofer LendelTorleif Härd

Abstract

Affibodies are a novel class of binding proteins selected from phagemid libraries of the Z domain from staphylococcal protein A. The Z(SPA-1) affibody was selected as a binder to protein A, and it binds the parental Z domain with micromolar affinity. In earlier work we determined the structure of the Z:Z(SPA-1) complex and noted that Z(SPA-1) in the free state exhibits several properties characteristic of a molten globule. Here we present a more detailed biophysical investigation of Z(SPA-1) and four Z(SPA-1) mutants with the objective to understand these properties. The characterization includes thermal and chemical denaturation profiles, ANS binding assays, size exclusion chromatography, isothermal titration calorimetry, and an investigation of structure and dynamics by NMR. The NMR characterization of Z(SPA-1) was facilitated by the finding that trimethylamine N-oxide (TMAO) stabilizes the molten globule conformation in favor of the fully unfolded state. All data taken together lead us to conclude the following: (1) The topology of the molten globule conformation of free Z(SPA-1) is similar to that of the fully folded structure in the Z-bound state; (2) the extensive mutations in helices 1 and 2 destabilize these without aff...Continue Reading

References

Feb 1, 1987·Protein Engineering·B NilssonM Uhlén
Aug 5, 1987·Journal of Molecular Biology·S MillerC Chothia
May 5, 1982·Journal of Molecular Biology·J Kyte, R F Doolittle
Jul 3, 1995·Proceedings of the National Academy of Sciences of the United States of America·H XiongM H Hecht
Jan 1, 1994·Nature Structural Biology·C RedfieldC M Dobson
Feb 1, 1995·Current Opinion in Structural Biology·O B Ptitsyn
Jul 1, 1994·Current Biology : CB·C M Dobson
Nov 1, 1995·Journal of Biomolecular NMR·F DelaglioA Bax
Feb 1, 1996·Journal of Molecular Graphics·R KoradiK Wüthrich
Aug 1, 1997·Nature Structural Biology·B A SchulmanC Redfield
Aug 1, 1997·Current Opinion in Structural Biology·P A Nygren, M Uhlén
Mar 28, 1998·The Journal of Biological Chemistry·I Baskakov, D W Bolen
May 7, 1999·Immunotechnology : an International Journal of Immunological Engineering·M HanssonP A Nygren
May 14, 1999·Current Biology : CB·C Redfield
Nov 11, 1999·Protein Engineering·E GunneriussonP Nygren
Mar 1, 2000·Trends in Biochemical Sciences·C CombetG Deléage
May 8, 2001·Annual Review of Biophysics and Biomolecular Structure·A G Palmer
Aug 15, 2001·Journal of Molecular Biology·D W Bolen, I V Baskakov
Jun 6, 2002·European Journal of Biochemistry·Jenny RönnmarkPer-Ake Nygren
Jul 27, 2002·Protein Science : a Publication of the Protein Society·Scott A Celinski, J Martin Scholtz
Dec 21, 2002·Protein Science : a Publication of the Protein Society·Yinan WeiMichael H Hecht
Jan 11, 2003·Journal of Biomolecular NMR·Christofer LendelTorleif Härd
Feb 21, 2003·Proceedings of the National Academy of Sciences of the United States of America·Elisabet WahlbergTorleif Härd
Jul 24, 2003·Biotechnology and Applied Biochemistry·Ronny FalkStefan Ståhl
Dec 25, 2003·Journal of the American Chemical Society·Wei Yuan YangMartin Gruebele

❮ Previous
Next ❯

Citations

Jul 4, 2009·Analytical Chemistry·Erin M JohnsonVicki H Wysocki
Aug 9, 2013·Protein Engineering, Design & Selection : PEDS·Malin LindborgTorleif Härd
Mar 17, 2010·Journal of Molecular Biology·Joachim FeldwischLars Abrahmsén
Aug 2, 2012·Protein Science : a Publication of the Protein Society·Adam W BarbJames H Prestegard
Jul 12, 2005·Current Opinion in Biotechnology·H Kaspar Binz, Andreas Plückthun
Nov 13, 2013·Biophysical Chemistry·Ryan D Macdonald, Mazdak Khajehpour
Jul 12, 2017·Bioconjugate Chemistry·Nika Kruljec, Tomaž Bratkovič
Oct 13, 2005·Journal of Molecular Recognition : JMR·Abdessamad Ababou, John E Ladbury
Sep 18, 2009·The Journal of Biological Chemistry·Lotta HolmMark Howarth
Jun 8, 2006·Journal of the American Chemical Society·Elisabet Wahlberg, Torleif Härd
Sep 10, 2004·Journal of the American Chemical Society·Vildan Dincbas-RenqvistTorleif Härd

❮ Previous
Next ❯

Related Concepts

Related Feeds

Bacteriophage: Phage Therapy

Phage therapy uses bacterial viruses (bacteriophages) to treat bacterial infections and is widely being recognized as an alternative to antibiotics. Here is the latest research.