The L chains from rabbit y-globulins can be separated by gel filtration into two populations (L1 and L2) differing markedly in their structural and antigenic proporties. The L l and L2 chains from pooled IgG have different amino-acid composition and half-cystine content. In addition, each population can be further resolved in two subfractions called L l a and Llb, L2a and L2b, respectively. The b allotypic markers are located exclusively in L2 chains.These results indicate that the L chains from rabbit IgG are constituted by two populations of different genetic origin.L chains from immunoglobulins can be classified in a restricted number of antigenic types, characterized by a constant C-terminal region. Two classes of L chains analogous to the x and A-chains previously described in man [l] et al. [6] have reported that L chains of rabbit y-globulins can be separated into two fractions. More recently, the characterization and the separation of two antigenically distinct groups of rabbit L chains has been achieved in our laboratory [7,8], and independently by slightly different methods by Mage et al. [9] using in both cases the procedure first described by Frangk and Nezlin [lo]. I n the present paper, we describe the separation of normal rabbit L chains into two populations differing in their antigenic and structural properties. MATERIAL AND METHODS Isolation Column Fractionation ProcedureThe heavy (H) and light (L) chains from IgG mildly reduced with 2.5 mM dithiothreitol and alkylated (refered to as mildly reduced and alkylated IgG) were separated by gel filtration on a column of Sephadex G-100 (3 x 140 cm) in 6 M urea, 0.05 M formic acid [lo].Each fraction was concentrated and purified by a second gel filtration under the same conditions. Immunological TestsAntiallotype sera were prepared by injecting Proteus vulgaris cells coated with antibody according to Dubiski et al. [13]. Sera and isolated polypeptide chains were tested against appropriate antisera by double diffusion analysis in Ouchterlony plates using a concentration of 1 01, agar in 0.05 M veronal buffer pH 8.6 [14]. The quantity of immunological precipitate of the L chains with the allotypic antisera was measured by simple radial immunodiffusion according to Mancini et al. [15].
Proglumide, a derivative of glutaramic acid which has been used for many years in the treatment of ulcers, was found to inhibit the degradation of the C-terminal octapeptide (CCK-8) of cholecystokinin (CCK) by brain enzymes. Another antagonist of CCK-gastrin receptors, dibutyryl cyclic GMP had no effect. The effect of proglumide was not specific for CCK peptides since the cleavage of the N-terminal tyrosine residue of leucine-enkephalin by brain enzymes was also inhibited. The inhibition did not involve CCK receptors as it was observed for the degradation of CCK-8 by cytosolic synaptosomal enzymes as well as for the cleavage of the peptide by purified synaptic membranes. The inhibition exerted by proglumide on the degradation of unsulfated CCK-8 by synaptic membranes displayed a non-competitive character with an apparent Ki value of 1.3 mM. Two non-competitive K, values (0.6 and 2.0 mM) were found for the inhibition of the cleavage of leu-enkephalin. Proglumide inhibited the degradation of CCK-8 by brain enzymes but was ineifective towards the cleavage of this peptide by pancreatic plasma membranes. Comparison of the structure of proglumide with aminopeptidase inhibitors shows some similarities between these molecules and suggests that this compound might act through the same mechanism on brain CCK-8 and leu-enkephalin degrading aminopeptidases.
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