[Hemoglobin alpha chain isoelectric point modification under the action of urea, sodium cyanate, succinic anhydride or diethylene triamine pentaacetic acid anhydride].

A monomeric protein, the hemoglobin alpha chain, was used to compare four protocols for conjugation with diethylene triamine pentaacetic (DTPA) anhydride. Carbamylation and succinylation were also performed. The isoelectric point (pI) was 7.7 for the native protein versus only 5.5 to 7.3 for the five carbamylated derivatives and 4.0 to 7.0 for the six succinylated derivatives. With carbamylation or succinylation, increasing the molar ratio (agent/protein) was associated with a gradual downward pI shift producing trains of bands. This phenomenon did not occur with DTPA conjugation, whose results varied with the method used; only one derivate (pI 6.7) was produced by all four methods, and multiple fine bands with pH values in the vicinity of 3.6 were seen. For the protein, the pI shift varied with the number of groups inserted on the primary amine residues. Also, the shift was larger if the inserted groups carried electrically-charged moieties.