Recombinant M-, B- and MB-type isozymes of human phosphoglyceric acid mutase: their large-scale production and preparation of polyclonal antibodies specific to M- and B-type isozymes

Abstract

Human phosphoglyceric acid mutase is a dimer comprising M-, B- and MB-type isozymes composed from the combination of the muscle-specific (M) and non-muscle-specific (B) subunits. Human DNAs coding M and B subunits were, respectively, reconstructed at their 5' regions without changing amino acid sequences, and expressed directly in Escherichia coli under the control of the trp promoter. M- and B-type isozymes were over-produced in the bacterial cytoplasm as soluble, active forms, which have been purified and characterized. MB-type was synthesized in vitro by recombining M- and B-type. All three recombinant isozymes thus obtained showed the same properties as the naturally-occurring ones with respect to the properties tested. Polyclonal IgGs specific to the M-type, B-type and MB-type were prepared from rabbits immunized with M- and B-type, using columns bound with M- and B-type. A method for the immunoassay of MB-type which is specifically present in cardiac muscle, is now under development.