Isolation of acute-phase proteins from plasma for determination of fractional synthesis rates by a stable isotope tracer technique.
Because of the unavailability of convenient tracer methods, there is no information on the kinetic changes responsible for the increased or decreased pool sizes of the acute-phase proteins (APPs) during the metabolic response to trauma and infections. We have developed a stable isotope tracer method to measure the synthesis rates of the APPs transferrin, haptoglobin, and alpha1-antitrypsin. The proteins were isolated from plasma by either direct or sequential immunoprecipitation and purified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. With the exception of haptoglobin, the major band of each protein and its standard ran to a position corresponding to its molecular mass. The major band of haptoglobin and its standard ran to a position corresponding to a molecular mass of slightly less than 44 kDa, which corresponds to the haptoglobin beta chain, molecular mass 42.5 kDa. To measure the fractional synthesis rates (FSRs) of these proteins, three children were infused with 2H3-leucine for 8 h, and the amount of 2H3-leucine incorporated into the proteins was measured by gas chromatography-mass spectrometry. The plateau isotopic enrichment of very low density lipoprotein-apoB-100-bound leucine was used to estimate the isotopic enrichment of the hepatic protein synthetic precursor pool. FSRs (%/day) of the proteins were haptoglobin, 50 +/- 8; transferrin, 19 +/- 1.1; and "alpha"1-antitrypsin, 10 +/- 1. 5.
Jahoor
F
F
USDA/ARS Children's Nutrition Research Center, Baylor College of Medicine, Houston, Texas, 77030, USA.
Sivakumar
B
B
Del Rosario
M
M
Frazer
E M
EM
D000209:Acute-Phase Proteins / Q000032:analysis* / Q000378:metabolism; D000328:Adult; D002675:Child, Preschool; D006801:Humans; D007930:Leucine / Q000378:metabolism; D008297:Male; D011233:Precipitin Tests; D011502:Protein-Energy Malnutrition / Q000097:blood; D014316:Tritium
D000209:Acute-Phase Proteins; D014316:Tritium; D007930:Leucine