FRACTIONAL SYNTHESIS RATES OF RETINOL-BINDING PROTEIN, TRANSTHYRETIN,AND A NEW PEPTIDE MEASURED BY STABLE-ISOTOPE TECHNIQUES IN NEONATAL PIGS

Our objective was to develop a stable isotopic method to measure the s ynthesis rates of retinol-binding protein (RBP) and transthyretin (TTR ). Both proteins were isolated from human and pig plasma by sequential immunoprecipitation and purified by SDS-polyacrylamide gel electropho resis under denaturing conditions. Both human and pig anti RBP precipi tates contained a peptide (TTR2) that had a molecular mass that was si milar but not identical to that of TTR subunit. The N-terminal amino a cid sequence of porcine TTR2 was highly but not completely homologous with porcine TTR. Human TTR2 showed no homology with TTR but was compl etely ho mologous with an internal sequence of human fibrinogen alpha chain. To measure the fractional rates of synthesis (FRS) of these pep tides, six infant pigs were infused with [H-2(3)]leucine at a constant rate for 6 h, and the amount of [H-2(3)]leucine incorporated into the proteins was measured by negative chemical ionization gas chromatogra phy-mass spectrometry. The plateau isotope ratio of plasma very low de nsity lipoprotein apoB-100-bound leucine was used to estimate the isot opic enrichment of hepatic protein synthetic precursor pool. The mean FRS (% h +/- S.E.) of TTR (1.97 +/- 0.13) and RBP (3.89 +/- 0.07) were significantly different. The FRS of TTR2 was low (0.31 +/- 0.19) rela tive to that of RBP and TTR. Thus, three different peptides with diffe rent turnover rates seem to be involved in the transport of retinol.