Development of crosslink-type tag-probe pairs for fluorescent imaging of proteins.
Useful methods of protein labeling via functional peptide tags have been developed in the field of proteome and chemical biology. New tag-probe pairs based on leucine zipper peptides for labeling target proteins are described. These consist of an α-helical probe peptide with an environmental-sensitive fluorescent dye and two antiparallel α-helical tag peptides, and may be crosslinked, from the Cys residue of the tag peptide to the N(α)-chloroacetyl group of the probe peptide. Binding of the probe peptide to the tag peptides results in movement of the fluorophore from a hydrophilic to a hydrophobic environment inside the leucine zipper structure, causing a dramatic fluorescent change, mediated by the binding of the two peptides. As a spacer between the N(α)-chloroacetyl group and the original probe sequence, a single Gly residue was the most suitable among 0-2 Gly residues. Crosslinking leads to superior fluorescence response, binding affinity, and chemical stability. These ZIP tag-probe pairs are useful for labeling and fluorescent imaging of proteins.
Nomura
Wataru
W
Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental University, Chiyoda-ku, Tokyo 101-0062, Japan.
Mino
Tomoaki
T
Narumi
Tetsuo
T
Ohashi
Nami
N
Masuda
Akemi
A
Hashimoto
Chie
C
Tsutsumi
Hiroshi
H
Tamamura
Hirokazu
H
D005456:Fluorescent Dyes / Q000737:chemistry*; D016350:Leucine Zippers; D015335:Molecular Probes / Q000138:chemical synthesis / Q000737:chemistry*; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry*; D017433:Protein Structure, Secondary; D011506:Proteins / Q000737:chemistry*
D005456:Fluorescent Dyes; D015335:Molecular Probes; D010455:Peptides; D011506:Proteins