Search

This report describes the characterization of a novel rat apolipoprotein, which, as partial sequencing suggests, does not correspond to any described protein. The protein (termed PX) has an estimated molecular mass of 19.5 kDa and pI in the range 5.5–5.8. Monoclonal antibodies were obtained against protein PX and results on distribution among rat lipoproteins show it to be associated mainly with high-density lipoproteins (HDL), but also with VLDL. Immunoaffinity chromatography of total HDL shows protein PX to be included in a distinct lipoprotein particle, particularly enriched in free cholesterol, with which only traces of other apolipoproteins are associated. Immunologically crossreacting entities are found in the plasma of several species, including man. Retention of the epitope carried by the protein PX would suggest that it is of particular structural or functional importance. It remains to be established whether its function is associated with lipid metabolism.

Two peptides, NA1 and NA2, which we previously suggested to be associated with high density lipoproteins (HDLs), have been purified. Polyclonal antibodies against each peptide and a monoclonal antibody against NA2 have been used to further characterize them and their association with HDL. Immunoblotting studies revealed that the peptides form a complex of molecular mass of approximately 80 kd. Agarose gel filtration showed coelution of NA1/NA2 and apolipoprotein (apo) A-I, the structural protein of HDL. This was confirmed by fast protein liquid chromatography, which further indicated that up to 60% of NA1/NA2 was located within the lower density range of the HDL spectrum. Complementary studies with anti-apo A-I immunoaffinity columns provided evidence that at least 40% of NA1/NA2 was associated with HDL, an association easily disrupted by ultracentrifugal manipulation. Finally, partial amino acid sequences showed virtually complete homology with a recently identified protein, SP-40,40, or cytolysis inhibitor. The protein is suggested to have a powerful inhibitory effect on complement-mediated cell lysis. Our results could thus furnish an explanation for the previously observed modulating influence of HDL on complement activity.