Dynamic interfacial properties of human apolipoproteins A-IV and B-17 at the air/water and oil/water interface

Abstract

Viscoelastic behavior of proteins at interfaces is a critical determinant of their ability to stabilize emulsions. We therefore used air bubble surfactometry and drop volume tensiometry to examine the dynamic interfacial properties of two plasma apolipoproteins involved in chylomicron assembly: apolipoprotein A-IV and apolipoprotein B-17, a recombinant, truncated apolipoprotein B. At the air/ water interface apolipoproteins A-IV and B-17 displayed wide area-tension loops with positive phase angles indicative of viscoelastic behavior, and suggesting that they undergo rate-dependent changes in surface conformation in response to changes in interfacial area. At the triolein/ water interface apolipoprotein A-IV displayed maximal surface activity only at long interface ages, with an adsorption rate constant of 1.0 ؋ 10 ؊ 3 sec ؊ 1 , whereas apolipoprotein B-17 lowered interfacial tension even at the shortest interface ages, with an adsorption rate constant of 9.3 ؋ 10 ؊ 3 sec ؊ 1 . Apolipoprotein A-IV displayed an expanded conformation at the air/water interface and a biphasic compression isotherm, suggesting that its hydrophilic amphipathic helices move in and out of the interface in response to changes in surface pressure. We conclude that apolipoproteins A-IV and B-17 display a combination of interfacial activity and elasticity particularly suited to stabilizing the surface of expanding triglyceride-rich particles.

Vesicle-binding properties of wild-type and cysteine mutant forms of α1 domain of apolipoprotein B

“…Herscovitz et al (24) demonstrated that wild-type apoB-17 is capable of forming recombinant discoidal lipoproteins on incubation with DMPC, a capability also observed for apoB-17F (25). Although the studies performed above indicated that disulfide bond mutations do not affect vesicle binding, we explored the possibility that these mutations may affect the efficiency by which apoB-17F converts DMPC MLV into recombinant lipoproteins.…”

“…Two micrograms of apoB-17F purified from baculovirusinfected Sf9 cells (25,28) or ϳ 3 ϫ 10 7 phosphorimager units of [ 35 S]apoB-17F from McArdle-RH7777 cells was incubated for 10 min at the indicated temperatures in the presence or absence of dithiothreitol (DTT) (final volume of 20 l). Free sulfhydryls were alkylated by adjusting the reaction to 100 mM iodoacetamide (IAA) and incubating at 37 Њ C for 20 min.…”

“…McArdle-RH7777 cells were stably transfected with a plasmid encoding the amino-terminal 17% of apoB appended with a C-terminal eight-amino acid FLAG epitope tag (apoB-17F) (25). Cells were radiolabeled for 24 h with [ 35 S]Met/Cys and the concentrated medium was incubated with (ϩ) or without (Ϫ) DMPC MLV for 20 h. After adjusting the density to 1.25 g/ml with KBr, samples were subjected to density gradient centrifugation.…”

“…To explore the role of native disulfide bonding in the binding of apoB-17F to DMPC vesicles, conditions were developed to achieve partial and complete chemical reduction. ApoB-17F immunoaffinity purified from recombinant baculovirus-infected Sf9 cell supernatant (25,28) was incubated in the absence (Ϫ) or presence (ϩ) of 50 mM DTT at the indicated temperatures (Fig. 2).…”

Effect of apolipoprotein A-IV genotype and dietary fat on cholesterol absorption in humans

“…This suggests that apoA-IV plays a specific role in the process of chylomicron assembly. Moreover, human apoA-IV displays unique conformational flexibility and elasticity at lipid/water interfaces (49), properties that are critical in stabilizing oil/water emulsion particles (50). Thus, by controlling molecular area at the expanding chylomicron surface during core lipidation, apoA-IV could modulate the influx of free cholesterol to the particle surface, thereby regulating particle cholesterol composition, and, ultimately, the efficiency of cholesterol absorption.…”