Primary structure of the bovine analogues to human apolipoproteins CII and CIII

Abstract

Two major isoforms of the bovine analogue to human apolipoprotein (apo) CII were purified from plasma. They were both as effective as human apo CII in activating lipoprotein lipase. Amino acid sequencing revealed that one form contained 79 amino acid residues, and corresponded to human pro apo CII. The other form lacked the first six residues at its N-terminus. This was apparently due to cleavage of the -Gln-Asp-linkage in the sequence H2N-Ala-His-Val-Pro-Gln-Gln-Asp-Glu-, analogous to cleavages described for human apo A1 and apo CII.Previous studies with human apo CII have shown that the ability to activate lipoprotein lipase resides in the C-terminal third of the molecule. This was highly conserved in the bovine analogue: of the 30 last residues, 21 are identical. Five residues in this part of human apo CII have been reported to be essential for activation of lipoprotein lipase. Only one of these, Tyr63, is present in the bovine sequence. The bovine structure contains a threonine at position 61, instead of serine in the human, and the four last residues are -Ser-Gly-Lys-Asp instead of the allegedly necessary -Lys-Gly-Glu-Glu. Three differently sialylated isoforms of the bovine analogue to human apolipoprotein CIII were also isolated and partially sequenced. All three lacked the first three N-terminal residues as compared to sequences from other species (man, dog and rat). Sequence differences were more pronounced at the ends than in the central parts of the apo CIII molecules.Transport of triacylglycerol in lipoproteins is a major pathway for energy transfer between tissues. Triacylglycerol-rich lipoproteins are formed in the intestine (chylomicra) and in the liver (very-low-density-lipoproteins). Their metabolism involves two major steps. Most of the triacylglycerols are unloaded by lipoprotein lipase at the vascular endothelium in peripheral tissues [ 11. The resulting remnant lipoproteins are then removed from the circulation mainly by the liver [2 -41. Apolipoproteins (apo) CII and CHI appear to be involved in the regulation of both steps. These proteins have flexible conformations [5] that allow them to bind reversibly to lipid/ water interfaces and to transfer rapidly between lipoprotein particles. When triacylglycerol-rich lipoproteins enter plasma they take up apo CII and CHI; when the particles are degraded by lipoprotein lipase the C apolipoproteins transfer back to other lipoproteins [6, 71. For hydrolysis by lipoprotein lipase the lipoproteins must carry apo CII, an activator for the enzyme [8, 91. In the absence of apo CII, triacylglycerol transport is severely impeded and gross hypertriglyceridemia ensues [lo]. Apo CIII has been shown to impede the interaction of lipoproteins with the remnant receptor, thus preventing their uptake by the liver until the lipase reaction has reduced them sufficiently in size so that most of the C apoproteins have been shed from the particles [4,11, 121. In addition, apo CIII has been shown to inhibit the action of lipoprotein lipase and the Human apo CII is a 79-a...

Apolipoprotein CII from Chicken (Gallus domesticus)

“…None of these Glu residues or a functional equivalent are present in chicken apoCII, or in the dog (10), the guinea pig (15), or the bovine apoCII proteins (11). Furthermore, one of the proposed Lys residues is lacking in guinea pig LPL even though this enzyme is activated by human apoCII (35).…”

“…Sequence homologies in apoCII. Sequence data for apoCII from mouse (14), rat (13), human (8), monkey (9), dog (10), sheep (12), cow (11), and guinea pig (15) are aligned with chicken apoCII. Negative numbers are used for amino acid residues in the signal peptides.…”

“…The activating material floated with the lipid emulsion on centrifugation. Thus, the lipid-binding proteins from chicken plasma could be isolated by the method that had been previously used for purification of apoCII from several other species (11,13,15). After delipidation, the precipitated proteins were run on a column of Sephadex G-75 (Fig.…”

“…The amino acid sequences are known for apoCII from eight mammalian species: human (8), monkey (9), dog (10), cow (11), sheep (12), rat (13), mouse (14), and guinea pig (15). The gene structures are known for human (16) and mouse apoCII (14).…”

Lipoprotein lipase activity and interactions studied in human plasma by isothermal titration calorimetry

“…Full-length human ANGPTL3, expressed in Sf 21 cells, was obtained from R&D Systems (USA). apoC-III 0 was purified from human plasma (21). Human apoC-II and human apoA-V were expressed in E. coli and purified as described (22,23).…”