Journal of Experimental Medicine, Vol 170, 811-825, Copyright © 1989 by Rockefeller University Press
Isolation and characterization of a distinct immunoregulatory isoform of alpha-fetoprotein produced by the normal fetus
NS van Oers, BL Cohen and RA Murgita
Department of Microbiology and Immunology, McGill University, Montreal, Quebec, Canada.
In this report, we examine the functional significance of the molecular
microheterogeneity of alpha-fetoprotein (AFP). In doing so, we have taken
the direct approach of purifying the naturally occurring isomeric forms of
fetal-derived AFP using a preparative anion exchange column linked to an
automated fast protein liquid chromatography (FPLC) system followed by
parallel testing of each isolated molecular variant for in vitro
immunoregulatory activity. The data obtained demonstrate the presence of
seven distinct variants of AFP as defined by their retention volumes on
FPLC elution profiles, by their pIs on analytical IEF gels, and by Western
blot analysis. Molecular mass determination by SDS-PAGE showed each isomer
to be equivalent in size to 69,000-dalton native unfractionated AFP
molecules. All the immunosuppressive activity of AFP was localized to a
single variant representing only 6% of the total composition of native AFP.
The immunoregulating isomer termed AFP- 1 was the least acidic of the seven
isolated variants with a pI of 5.1 and displayed a sialic acid content of 1
mol/mol of protein. The inhibitory activity of AFP-1 could be readily
measured on T cell- dependent antibody synthesis, Con A-induced stimulation
of Lyt-1+23- thymocyte DNA synthesis, and lymphokine-activated NK cell
activity. All other isomers were without effect in these test systems. The
immunosuppressive AFP-1 isomer also displayed the strongest growth-
promoting influence on cultured bone marrow lymphocytes. There was no
correlation between functional activity and degree of expression of sialic
acid residues on the AFP molecules. These findings demonstrate that the
immunoregulating function of AFP is confined to a distinct and relatively
small subpopulation of native AFP molecules and should therefore contribute
to the resolution of outstanding questions regarding the structure/function
relationship of this onco-fetal glycoprotein.
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