Myeloma proteins with anti-DNP activity

The method is also capable of differentiating among different antibody populations with the same specificity. Mouse anti-Dnp antibodies elicited by immunization with Dnp derivatives induce circular dichroic patterns, upon interaction with e-Dnp-lysine, that are strikingly different from those induced by MOPC 315, a mouse IgA myeloma protein with anti-Dnp activity (38). It was concluded that the combining site of protein 315 must differ in its fine structure from that of the majority of induced mouse anti-Dnp antibodies. 

With the exception of a myeloma protein with anti-Dnp activity, preparations used for these studies have been specifically purified antihapten antibodies, which are heterogeneous. The velocity constants reported, as well as equilibrium constants, are therefore average values. 

Table 2.5 summarizes data obtained with a number of haptens and their corresponding antibodies. Except for protein 315, a mouse myeloma protein with anti-Dnp activity, specifically purified rabbit antibodies were used. The data on reaction rates are expressed in terms of a second-order velocity constant, k, where the forward reaction velocity is V = A i2(A) (L) v is in mole/liter/second (A) and (L) are the molar concentrations of free (unoccupied) antibody combining sites and of ligand, respectively. The units of ki2 are 1/mole/second (M sec ). In measuring rates it is essential to have a method for detection of the product at a low concentration, since the concentrations of reactants must be sufficiently small so that the velocity of the reaction is not too rapid to measure. 

All preparations are specifically purified rabbit antibodies except for protein 315, which is a mouse myeloma protein with anti-Dnp activity. 

Protein 460, another mouse myeloma protein with anti-Dnp activity, was also affinity labeled by compound BADE (136). The label was found on the L chain, as in protein 315, but attached to a lysyl rather than a tyrosyl side chain. The locus of attachment was in a hypervariable region, at position 54. 

Specificity of a Human Myeloma Protein with Anti-Dnp Activity... 

Rather extensive studies have been carried out on mouse (BALB/c) myeloma proteins with specificity for the Dnp group, phosphorylcholine or a(l 3)dextran. One protein with anti-Dnp activity, designated MOPC 315, was elicited by M. Potter and its properties were described by Eisen et al. (87). The activity was discovered by screening a large number of BALB/c myeloma proteins for their reaction with the Dnp hapten by the method of spectral shift analysis that had already been applied to screen human myeloma sera. Protein 315 has a binding affinity for e-Dnp-L-lysine of approximately 10 M and a very similar affinity for -Dnp-6-aminocaproate. The H chains are of the IgA class but the L chains have an unusual amino acid sequence which resembles that of normal X chains, but differs with respect to approximately 27 residues in the C-terminal half (Cl domain). On the basis of this comparison, Schu-lenburg et al. (88) concluded that there are at least two subtypes of... 

The diazo group becomes activated in the presence of light. The usefulness of this reagent is based, in part, on the fact that certain mouse myeloma proteins (MOPC 315 and 460) with anti-Dnp activity also react with menadione. 

This immunologic cross-reaction, whose chemical basis is not readily apparent, is also observed with rabbit anti-Dnp antibodies. In any event, it has been possible to aflinity label myeloma protein 460, which has anti-Dnp activity, with H-MenDK. Approximately two-thirds of the active sites of the myeloma protein were covalently labeled about 85% of the label was present in the H chain (130). 

Related findings were reported by Eisen and his collaborators, who showed that mouse myeloma protein, MOPC 315, which has anti-Dnp activity, also combines weakly with 5-acetyluracil, caffeine, and riboflavin, and somewhat more strongly with menadione (2-methyl-1,4-naphthaquinone) (105). None of these compounds is closely related structurally to the dinitrophenyl group. Cross-reactions with 5-acetyluracil of anti-Dnp antibodies induced in some but not all rabbits and guinea pigs were also observed (106). 

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