Specific receptor sites for pyridoxal 5'-phosphate and pyridoxal 5'-deoxymethylenephosphonate at the alpha and beta NH2-terminal regions of hemoglobin.

hemoglobins have been prepared which are substituted with pyridoxal 5'-phosphate or pyridoxal 5'-deoxymethylenephosphonate at both NH2-terminal amino groups of either the alpha or the beta chains. 31P NMR titration curves of the 5' side chain show a substantial decrease in acid strength in hemoglobins pyridoxylated at the alpha chains and the reverse effect in the hemoglobins labeled in the beta chains. These changes can readily be explained by the interaction of the 5' side chains with surrounding residues which provide specific binding sites for the monoanion at the alpha chain termini and for the dianion in the case of the beta chains. Since the proportion of monoanion is at lest ten times greater for pyridoxal 5'-deoxymethylenephosphonate than for pyridoxal 5'-phosphate at neutral pH, the phosphonate reacts preferentially with the alpha chain NH2-terminal amino groups and the phosphate with the beta chain ones.