Apolipoprotein C-II (
apoC-II) is an exchangeable plasma
apolipoprotein and an endogenous activator of
lipoprotein lipase (LpL). Genetic deficiencies of
apoC-II and overexpression of
apoC-II in transgenic mice are both associated with severe
hyperlipidemia, indicating a complex role for
apoC-II in the regulation of blood
lipid levels.
ApoC-II exerts no effect on the activity of LpL for soluble substrates, suggesting that activation occurs via the formation of a
lipid-bound complex. We have synthesized a
peptide corresponding to
amino acid residues 39-62 of mature human
apoC-II. This
peptide does not bind to model
lipid surfaces but retains the ability to activate LpL. Conjugation of the fluorophore
7-nitrobenz-2-oxa-1,3-diazole (NBD) to the N-terminal alpha-amino group of apoC-II39-62 facilitated determination of the affinity of the
peptide for LpL using fluorescence anisotropy measurements. The dissociation constant describing this interaction was 0.23 microM, and was unchanged when LpL was
lipid-bound. Competitive binding studies showed that apoC-II39-62 and full-length
apoC-II exhibited the same affinity for LpL in aqueous
solution, whereas the affinity for full-length
apoC-II was increased at least 1 order of magnitude in the presence of
lipid. We suggest that while the binding of
apoC-II to the
lipid surface promotes the formation of a high-affinity complex of
apoC-II and LpL, activation occurs via direct helix-helix interactions between apoC-II39-62 and the loop covering the active site of LpL.