Bz. Packard et al., STRUCTURAL CHARACTERISTICS OF FLUOROPHORES THAT FORM INTRAMOLECULAR H-TYPE DIMERS IN A PROTEASE SUBSTRATE, JOURNAL OF PHYSICAL CHEMISTRY B, 101(25), 1997, pp. 5070-5074
Recently, we designed and synthesized a new class of profluorescent pr
otease substrates whose spectral properties fit the exciton model: mor
e specifically, spectra of these polypeptide which were doubly labeled
with rhodamines showed blue shifted absorption peaks and fluorescence
quenching, both indicators of H-dimer formation. In the work describe
d here NorFES, an undecapeptide which is cleaved by the serine proteas
e elastase, was homodoubly labeled on opposite sides of its cleavage s
ite with six fluorophores in order to identify structural elements of
dyes which influence intramolecular H-type dimer formation, Absorption
and fluorescence spectra of these six substrates obtained before and
after enzymatic cleavage indicate that the exciton band is strongest i
n the peptide doubly labeled with tetramethylrhodamine, followed by rh
odamine-X. and then (diethylamino)coumarin. In contrast, spectra of No
rFES homodoubly labeled with fluorescein, hydroxycoumarin, or pyrene d
o not exhibit exciton bands, These data suggest that factors significa
nt in H-type dimerization are as follows (in decreasing order): deloca
lized charge, symmetry. and magnitude of the lowest energy electronic
transition dipole. Surprisingly, in the group of fluorophores in this
study, no evidence for hydrophobic interactions as an important influe
nce was observed.