Influence of the mercury blocking reagent 2-mercaptoethanol on the spectroscopic properties of complexes formed between lysyltryptophyllysine and mercurated poly(uridylic acid).

Abstract

Optical detection of magnetic resonance (ODMR) studies are reported for complexes formed between the tripeptide Lys-Trp-Lys and poly(5-HgU), both in the absence and in the presence of the blocking reagent 2-mercaptoethanol (ME). Complexes formed both with and without ME show characteristics of a heavy atom effect: quenching of Trp fluorescence, enhancement of the Trp phosphorescence quantum yield, a drastic reduction of the Trp phosphorescence lifetime, and the appearance of a Trp magnitude of D + magnitude of E ODMR signal at ca. 4.2 GHz. Significant differences are found, however, in the photophysical properties of the complexes formed with and without ME. In the absence of ME, the Trp phosphorescence bands are broad, the 0,0 band is shifted to 415.3 nm, and the magnitude of D - magnitude of E and 2 magnitude of E ODMR transitions are broad and poorly resolved. These features are characteristic of an inhomogeneous Trp environment. In the presence of ME, the phosphorescence peaks are narrower, with the 0,0 band shifted to 411.6 nm. The magnitude of D - magnitude of E and 2 magnitude of E ODMR transitions are well resolved and shifted in frequency relative to the unblocked complex. These features point to a more homogeneous Trp environment in the presence of ME. UV difference spectra show hypochromicity in the poly(5-HgU) absorption band (indicating induced stacking) which occurs on binding of Lys-Trp-Lys with ME present, while in the absence of ME, hypochromicity occurs primarily in the Trp absorption bands. Reversal of these effects with added NaClO4 occurs in both cases, but higher ionic strength is required with ME present. These results are consistent with the formation of stacked complexes in the presence of ME, but with additional types of complexes in its absence. The additional complexes formed in the absence of ME do not contribute to stacking of poly-(5-HgU) and may involve direct binding of mercury to amines of Lys-Trp-Lys; binding occurs between Lys-Trp-Lys and the monomer 5-HgUTP in the absence of ME, but not when the Hg is blocked with ME.