Polybrominated diphenyl ethers (PBDEs) have been shown
to disrupt
thyroid hormone (TH) functions on experimental animals, and one of
the proposed disruption mechanisms is the competitive binding of PBDE
metabolites to TH transport proteins. In this report, a nonradioactive,
site-specific fluorescein–thyroxine (F–T4) conjugate
was designed and synthesized as a fluorescence probe to study the
binding interaction of hydroxylated PBDEs to thyroxine-binding globulin
(TBG) and transthyretin (TTR), two major TH transport proteins in
human plasma. Compared with free F–T4, the fluorescence intensity
of TTR-bound conjugate was enhanced by as much as 2-fold, and the
fluorescence polarization value of TBG-bound conjugate increased by
more than 20-fold. These changes provide signal modulation mechanisms
for F–T4 as a fluorescence probe. Based on fluorescence quantum
yield and lifetime measurements, the fluorescence intensity enhancement
was likely due to the elimination of intramolecular fluorescence quenching
of fluorescein by T4 after F–T4 was bound to TTR. In circular
dichroism and intrinsic tryptophan fluorescence measurements, F–T4
induced similar spectroscopic changes of the proteins as T4 did, suggesting
that F–T4 bound to the proteins at the T4 binding site. By
using F–T4 as the fluorescence probe in competitive binding
assays, 11 OH–PBDEs with different levels of bromination and
different hydroxylation positions were assessed for their binding
affinity with TBG and TTR, respectively. The results indicate that
the binding affinity generally increased with bromine number and OH
position also played an important role. 3-OH–BDE-47 and 3′-OH–BDE-154
bound to TTR and TBG even stronger, respectively, than T4. With rising
environmental level and high bioaccumulation capability, PBDEs have
the potential to disrupt thyroid homeostasis by competitive binding
with TH transport proteins.