A Highly Conserved Motif within the NH2-terminal Coiled-coil Domain of Angiopoietin-like Protein 4 Confers Its Inhibitory Effects on Lipoprotein Lipase by Disrupting the Enzyme Dimerization

Abstract

Lipoprotein lipase (LPL) is a principal enzyme responsible for the clearance of chylomicrons and very low density lipoproteins from the bloodstream. Two members of the Angptl (angiopoietin-like protein) family, namely Angptl3 and Angptl4, have been shown to inhibit LPL activity in vitro and in vivo. Here, we further investigated the structural basis underlying the LPL inhibition by Angptl3 and Angptl4. By multiple sequence alignment analysis, we have identified a highly conserved 12-amino acid consensus motif that is present within the coiled-coil domain (CCD) of both Angptl3 and Angptl4, but not other members of the Angptl family. Substitution of the three polar amino acid residues (His 46 , Gln 50 , and Gln 53 ) within this motif with alanine abolishes the inhibitory effect of Angptl4 on LPL in vitro and also abrogates the ability of Angptl4 to elevate plasma triglyceride levels in mice. The CCD of Angptl4 interacts with LPL and converts the catalytically active dimers of LPL to its inactive monomers, whereas the mutant protein with the three polar amino acids being replaced by alanine loses such a property. Furthermore, a synthetic peptide consisting of the 12-amino acid consensus motif is sufficient to inhibit LPL activity, although the potency is much lower than the recombinant CCD of Angptl4. In summary, our data suggest that the 12-amino acid consensus motif within the CCD of Angptl4, especially the three polar residues within this motif, is responsible for its interaction with and inhibition of LPL by blocking the enzyme dimerization.

Mechanisms involved in biological behavior changes associated with Angptl4 expression in colon cancer cell lines

“…It is composed of 406 amino acids and contains an amino-terminal coiled-coil domain and a carboxyl-terminal fibrinogen-like domain (3). The most studied functions of Angptl4 are its role in the regulation of angiogenesis, glucose metabolism and lipid metabolism (4)(5)(6).…”

Fatty Acids Bind Tightly to the N-terminal Domain of Angiopoietin-like Protein 4 and Modulate Its Interaction with Lipoprotein Lipase

“…This is supported by population-based studies that indicate that individuals carrying the E40K variant of Angptl4 have low plasma triglyceride levels (26). Furthermore, a synthetic peptide spanning residues 44 -55 of human Angptl4 was able to inhibit LPL, but the affinity of this peptide for LPL was much lower than that of full-length ccd-Angptl4 (25).…”

“…This was supported by fluorescence measurements, demonstrating that fatty acids hinder the accessibility of the quencher acrylamide to the single Trp residue of ccd-Angptl4. This residue (Trp-38) is located close to the putative binding site for LPL (24,25). It has been previously shown that ccd-Angptl4 induces inactivation of LPL by causing dissociation of dimeric LPL into inactive monomers (10).…”

“…The N-terminal domain of Angptl4 binds to LPL with high affinity, resulting in inactivation of the enzyme (10,23). It has been proposed that the LPL-binding region in Angptl4 is located between residues 38 and 52 (24,25). This is supported by population-based studies that indicate that individuals carrying the E40K variant of Angptl4 have low plasma triglyceride levels (26).…”

Nutritional regulation of adipose tissue lipoprotein lipase is blunted in insulin resistant rats

“…Subsequent studies employing transgenic mice overexpressing angptl4 specifically in white adipose tissue have shown that this protein significantly decreased postheparin plasma LPL activity [17]. Numerous studies have provided firm evidence that angptl4 irreversibly inhibits LPL activity which is at least partly accounted for by promoting the conversions of active LPL dimers into inactive monomers [18]. In rodents, angptl4 is most highly expressed in adipose tissue followed by the liver, intestine, heart, kidney and ovary [19].…”