Protein kinetics determined in vivo with a multiple-tracer, single-sample protocol: application to lactase synthesis

The American Journal of Physiology
M A DudleyP J Reeds

Abstract

Precise analysis of the kinetics of protein/enzyme turnover in vivo has been hampered by the need to obtain multiple tissue samples at different times during the course of a continuous tracer infusion. We hypothesized that the problem could be overcome by using an overlapping (i.e., staggered) infusion of multiple stable amino acid isotopomers, which would take the place of multiple tissue samples. We have measured, in pigs, the in vivo synthesis rates of precursor (rapidly turning over) and mature (slowly turning over) polypeptides of lactase phlorizin hydrolase (LPH), a model for glycoprotein synthesis, by using an overlapping infusion of [2H3]leucine, [13C1]leucine, [13C1]phenylalanine, [2H5]phenylalanine, [13C6]phenylalanine, and [2H8]phenylalanine. Blood samples were collected at timed intervals, and the small intestine was collected at the end of the infusion. The tracer-to-tracee ratios of each isotopomer were measured in the plasma and jejunal free amino acid pools as well as in purified LPH polypeptides. These values were used to estimate kinetic parameters in vivo using a linear steady-state compartmental model. The fractional synthesis rates of the high-mannose, complex glycosylated and mature brush-border LPH polype...Continue Reading

References

Feb 15, 1979·The Biochemical Journal·M A McNurlanP J Garlick
Oct 1, 1992·Journal of Pediatric Gastroenterology and Nutrition·M A DudleyB L Nichols
Mar 1, 1992·The Journal of Nutrition·M A DudleyP J Reeds
Feb 1, 1992·Journal of Pediatric Gastroenterology and Nutrition·R J ShulmanS J Henning
Jun 1, 1971·Journal of Animal Science·R F ElliottH Fisher
Jul 1, 1984·The Biochemical Journal·E M DanielsenH Sjöström
Jul 1, 1994·Gastroenterology·L MaiuriS Auricchio
May 1, 1997·The Journal of Nutrition·M A DudleyP J Reeds
Sep 15, 1991·Proceedings of the National Academy of Sciences of the United States of America·H K BertholdP D Klein

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Citations

Sep 10, 2002·American Journal of Physiology. Endocrinology and Metabolism·Xiao-Jun ZhangRobert R Wolfe
Oct 22, 2010·The Journal of Physiology·Francesco AgostiniGianni Biolo
Mar 20, 2001·American Journal of Physiology. Gastrointestinal and Liver Physiology·M A DudleyP J Reeds
Jan 7, 2003·The Journal of Nutrition·Teresa A DavisMarta L Fiorotto

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