Changes of the main isoform of human apolipoprotein A-I following incubation of plasma

Abstract

This study was aimed to ascertain whether the more acidic isoforms of plasma apo A-I (A-I −1, and A-I −2) could originate in vitro from the main plasma isoform (A-I 0). Apo A-I isoforms were separated by two-dimensional gel electrophoresis before and after a prolonged incubation of serum or EDTA-plasma at 37°C. In incubated plasma there was a marked decrease of apo A-I 0 and a concomitant linear increase of apo A-I −1 + and A-I −2. The relative content of the latter raised from 22 ± 7% before the incubation to 60 ± 7% after 48 h of incubation. This conversion of A-I 0 was not inhibited by either pre-heating of plasma at 60°C for 1 h or the addition of protease inhibitors, EDTA and p-chloromercuriphenylsulfonic acid. The conversion of apo A-I 0 was not observed in isolated HDL, incubated either in the absence or in the presence of d < 1.063 g/ml lipoproteins and lipoprotein deficient plasma, nor in plasma which had been dialyzed before being incubated at 37°C. This suggests that plasma contains a low molecular weight factor capable of promoting the conversion of A-I 0. The increase of the relative content of apo A-I −1 and apo A-I −2 in incubated plasma was not due to glucosylation or carbamylation of A-I 0 as no radioactive glucose and urea were found to be bound to A-I. Since the conversion of apo A-I 0 was prevented by the addition of an antioxidant (butylated hydroxytoluene, BHT) to the incubated plasma it is conceivable that some product of lipid peroxidation renders apo A-I 0 more electronegative by reacting with some free amino groups of this peptide.