bc9b00786_si_001.pdf (1.14 MB)
Photocontrolled Reversible Binding between the Protein A‑Derived Z Domain and Immunoglobulin G
journal contribution
posted on 2020-02-25, 15:42 authored by Anders Myrhammar, Daniel Rosik, Amelie Eriksson KarlströmPhotoisomerization
of the trans and cis isomers of
azobenzene derivatives has been used to control the function
of biomolecules in a reversible and nondestructive manner. In this
study, affibody molecules, representing a class of small, helical
proteins that can be engineered for binding to a wide range of target
proteins, have been investigated by the incorporation of a photoswitchable
azobenzene derivative in the molecule. Three different Z domain variants
were produced by solid phase peptide synthesis and conjugated by thiol-directed
chemistry to an azobenzene-based photoswitch. The proteins were screened
for binding to and light elution from an IgG-sepharose affinity column.
One of the tested Z variants, ZC3, showed efficient binding
to the column and could be eluted by irradiation with light at 400
nm. In a reverse affinity chromatography assay, where the ZC3 variant was coupled to sepharose, human IgG1 could be captured to
the column and partially eluted by light. Further studies of the azobenzene-conjugated
ZC3 domain by surface plasmon resonance (SPR) confirmed
the high affinity binding to IgG, and circular dichroism (CD) spectroscopy
showed that the protein has a high α-helical secondary structure
content.
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Keywords
affinity bindingtarget proteinsSPRstructure contentazobenzene derivativeslight elutionZ C 3 variantazobenzene-based photoswitchphase peptide synthesisImmunoglobulin G Photoisomerizationα- helicalZ domain variantsazobenzene-conjugated Z C 3 domainIgG-sepharose affinity columnphotoswitchable azobenzenehelical proteinsIgG 1affibody moleculesZ variantscis isomerssurface plasmon resonancePhotocontrolled Reversible BindingZ C 3affinity chromatography assay400 nmthiol-directed chemistry
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