Protein residualizing labels facilitate localization of tissue sites of protein catabolism and the quantification of protein accumulation because of their prolonged intracellular retention of protein accumulation because of their prolonged intracellular retention times. Radioiodinated residualizing labels have been used to define the metabolism of a wide variety of proteins, but this has necessitated destructive analysis. Here we describe the implementation and validation of a novel 19F-containing residualizing label for protein, NN-dilactitol-3,5-bis(trifluoromethyl)benzylamine (DLBA), that permits the non-invasive assessment of protein accumulation and catabolism by n.m.r. spectroscopy in vivo. DLBA comprises a reporter molecule containing six equivalent 19F atoms. 19F is strongly n.m.r.-active, has 100% natural abundance, and is present in minimal background concentrations in soft tissues. We validated the use of DLBA as a protein-labelling compound by coupling to asialofetuin (ASF), a protein that is recognized exclusively by hepatic tissue via a saturable receptor-mediated process. Coupling of DLBA to ASF by reductive amination had no effect on the physiological receptor-mediated uptake of the protein in rat liver in vivo. The 19F-n.m.r. spectrum of DLBA exhibited a single peak that was subject to a small chemical-shift change and broadening after coupling to ASF. Pronase digestion of DLBA-ASF was performed to simulate intracellular degradation products, and resulted in a narrower set of resonances, with chemical shifts intermediate between those of uncoupled DLBA and DLBA-ASF. Intravenous administration of DLBA-ASF to rats followed by quantification of 19F in homogenates of liver tissue indicated that the half-life of residence time of degradation products from DLBA-ASF in liver was approx. 2 days. This intracellular half-life was comparable with that described for similar residualizing labels that contain radioiodide as a reporter. Similar results for the half-life of retention were obtained non-destructively and non-invasively in situ with the use of a whole-body radio-frequency antenna to acquire sequential spectra over 80 h after intravenous administration of DLBA-ASF. Quantification of these spectra demonstrated an initial accumulation of DLBA-ASF in liver followed by an expected gradual loss of 19F-labelled degradation products. The approach developed offers promise for the sequential and longitudinal characterization of metabolism of specific proteins in individual experimental animals and ultimately in human subjects.
Skip Nav Destination
Article navigation
December 1989
-
Cover Image
Cover Image
- PDF Icon PDF LinkFront Matter
- PDF Icon PDF LinkTable of Contents
- PDF Icon PDF LinkAdvertising
Research Article|
December 15 1989
Non-invasive detection of protein metabolism in vivo by n.m.r. spectroscopy. Application of a novel 19F-containing residualizing label
A Daugherty;
A Daugherty
*Department of Internal Medicine, Washington University School of Medicine, St Louis, MO 63110, U.S.A.
Search for other works by this author on:
N N Becker;
N N Becker
†Department of Chemistry, Washington University, St Louis, MO 63130, U.S.A.
Search for other works by this author on:
L A Scherrer;
L A Scherrer
†Department of Chemistry, Washington University, St Louis, MO 63130, U.S.A.
Search for other works by this author on:
B E Sobel;
B E Sobel
*Department of Internal Medicine, Washington University School of Medicine, St Louis, MO 63110, U.S.A.
Search for other works by this author on:
J J H Ackerman;
J J H Ackerman
*Department of Internal Medicine, Washington University School of Medicine, St Louis, MO 63110, U.S.A.
†Department of Chemistry, Washington University, St Louis, MO 63130, U.S.A.
Search for other works by this author on:
J W Baynes;
J W Baynes
‡Department of Chemistry, University of South Carolina, Columbia, SC 29208, U.S.A.
§Department of Medicine, University of South Carolina, Columbia, SC 29208, U.S.A.
Search for other works by this author on:
S R Thorpe
S R Thorpe
‡Department of Chemistry, University of South Carolina, Columbia, SC 29208, U.S.A.
Search for other works by this author on:
Publisher: Portland Press Ltd
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© 1989 London: The Biochemical Society
1989
Biochem J (1989) 264 (3): 829–835.
Citation
A Daugherty, N N Becker, L A Scherrer, B E Sobel, J J H Ackerman, J W Baynes, S R Thorpe; Non-invasive detection of protein metabolism in vivo by n.m.r. spectroscopy. Application of a novel 19F-containing residualizing label. Biochem J 15 December 1989; 264 (3): 829–835. doi: https://doi.org/10.1042/bj2640829
Download citation file:
Sign in
Don't already have an account? Register
Sign in to your personal account
You could not be signed in. Please check your email address / username and password and try again.
Could not validate captcha. Please try again.