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. Author manuscript; available in PMC: 2019 Dec 13.
Published in final edited form as: J Phys Chem B. 2018 Oct 10;122(49):11702–11720. doi: 10.1021/acs.jpcb.8b07768

Figure 1.

Figure 1.

Immobilization of dye-labeled α3D. (a) The donor (D), Alexa 488, is attached to the N-terminal unnatural amino acid (4-acetylphenylalanine), acceptor 1 (A1), Alexa 594, is attached to cysteine in the middle of helix 2 (residue 33), and acceptor 2 (A2) is attached to the C-terminal cysteine residue. Molecules are immobilized on a polyethylene glycol (PEG)-coated glass surface via a biotin-NeutrAvidin linkage. (b) After labeling D and A1 site-specifically, a cysteine residue is appended at the C-terminus of the protein using the sortase-mediated ligation. Then, A2 is attached to the C-terminal cysteine residue.