Long-chain fatty acid-binding to albumin: re-evaluation with directly measured concentrations.
In studies on uptake of fatty acids (FA) into organs, the unbound (or free) fatty acid fraction is commonly calculated from the concentration bound to albumin and from published binding constants. However, there is some dispute on the methods used for determining those binding constants. We developed a method allowing direct measurement of unbound FA by extending the previous studies of Svenson et al. [1] and Reed et al. [2]. Albumin was coupled to a solid phase (Sepharose 4B), loaded with FA and equilibrated with an aqueous solution. Laurate, palmitate and oleate concentrations in the aqueous phase were determined at different molar ratios of FA to albumin (r) and at different temperatures. FA albumin-binding constants (Ki) increase with chain length and decrease with temperature, in accordance with data obtained by others. However, the unbound concentrations measured are markedly lower than those obtained from binding constants, and the resulting Ki values markedly higher. This difference is presumed to result from (1) our direct measurement of unbound FA and (2) utilizing different more physiological conditions. Recalculating kinetic parameters from published FA uptake data, we found considerably different Km and Vmax values compared to the original data. Thus, the FA-binding characteristics measured in this study may influence the interpretation of FA uptake substantially.
Rose
H
H
Institute of Physiology, Med. Fac. RWTH Aachen, Germany.
Conventz
M
M
Fischer
Y
Y
Jüngling
E
E
Hennecke
T
T
Kammermeier
H
H
D000418:Albumins / Q000378:metabolism*; D002021:Buffers; D005227:Fatty Acids / Q000378:metabolism*; D005230:Fatty Acids, Nonesterified / Q000032:analysis*; D008433:Mathematics; D012685:Sepharose; D012710:Serum Albumin, Bovine / Q000378:metabolism; D013696:Temperature; D013816:Thermodynamics
D000418:Albumins; D002021:Buffers; D005227:Fatty Acids; D005230:Fatty Acids, Nonesterified; D012710:Serum Albumin, Bovine; D012685:Sepharose