Apolipoprotein A-IV has been isolated from four sources: human and rat lymph and plasma. Conformational properties of the rat and human apoA-IV in solution and denaturation changes induced by guanidine hydrochloride (Gnd · HCl) were studied using circular dichroic and fluorescence spectroscopy, and analytical sedimentation equilibrium ultracentrifugation. We have shown that both rat and human apoA-IV have similar secondary structure with negative maxima in the circular dichroic spectra at 222 nm and 207 nm. Furthermore, we have found no significant difference in the α-helical content of the apoA-IV from rat plasma (33%), rat lymph (37%), human plasma (35%), or human lymph (35%). Our denaturation studies with Gnd · HCl demonstrated reversibility and the fact that each apoA-IV had a tendency to self-associate in solution and the self-association could be disrupted by low concentrations of Gnd · HCl (≤ 0.4 M). Unfolding of the secondary structure of each apoA-IV occurred at higher concentrations of Gnd · HCl (midpoint ≤ 1.0 M). The apparent free energy of denaturation of the four apoA-IV proteins calculated from changes in the circular dichroic spectra upon addition of increasing concentrations of Gnd · HCl varied in a range from 3.0 to 4.2 kcal/mol. The fluorescence experiments revealed that apoA-IV from all sources had a maximum fluorescence emission at 342.5 nm, which shifted to the red region upon addition of increasing concentrations of Gnd · HCl. Fluorescence quenching of apoA-IV by neutral (acrylamide) and negatively charged (iodide ion) quenches demonstrated the increased accessibility of the tryptophanyl residues under the denaturing conditions. We conclude that there are no significant differences in the solution properties of apoA-IV isolated from rat plasma, rat lymph, human plasma, or human lymph and, that apoA-IV from both species is thermodynamically unstable in aqueous solution.
|Original language||English (US)|
|Number of pages||9|
|Journal||Journal of lipid research|
|State||Published - 1985|
ASJC Scopus subject areas
- Cell Biology