Journal of Lipid Research, Vol 35, 458-467, Copyright © 1994 by Lipid Research, Inc.
13C NMR studies of the binding of medium-chain fatty acids to human serum albumin
MA Kenyon and JA Hamilton
Department of Biophysics, Housman Research Center, Boston University Medical School, MA 02118-2394.
Binding of the medium-chain fatty acids (MCFA), octanoic (OCT) and decanoic
(DEC) acid, to human serum albumin (HSA) has been studied by 13C NMR
spectroscopy. NMR spectra at 35 degrees C showed an apparently homogeneous
binding environment (a single, narrow resonance for the 13C- enriched
carboxyl carbon) at different mole ratios and pH values. Changes in the
chemical shift of this peak with mole ratio and protein concentration
demonstrated rapid equilibration (< or = msec) of bound and unbound MCFA
and permitted a direct quantitation of bound/unbound MCFA. Spectra of
OCT/HSA mixtures at 6 degrees C revealed at least three distinct binding
sites that fill sequentially. The observed heterogeneity of binding at low
temperature, compared to 35 degrees C, is attributed to a slower exchange
rate of OCT between binding sites. The highest affinity sites for both OCT
and DEC have properties similar to those of binding sites for longer-chain
fatty acids, such as the close proximity of the fatty acid carboxylate to
basic amino acid residue(s). Interestingly, chemical shift data showed that
the first mole of OCT and DEC either bind differently to the same site or
bind to different sites on HSA. The rapid desorption of MCFA from HSA
binding sites has implications for dietary regimens with medium chain
triglycerols.
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