DELTA-AMINOLEVULINATE INCREASES HEME SATURATION AND YIELD OF HUMAN CYSTATHIONINE BETA-SYNTHASE EXPRESSED IN ESCHERICHIA-COLI

We recently expressed human cystathionine beta-synthase (CBS) in Esche richia coli and purified it to homogeneity, We showed that CBS require s heme in addition to pyridoxal 5'-phosphate for its function, Previou sly, CES, only about 20% saturated with heme, was purified from transf ormed bacteria, In the present study, we supplemented the bacteria wit h 0.3 mM delta-aminolevulinate (delta ALA), a precursor of heme, While growth of the bacteria did not change, a 50-fold elevation of the hem e content per milligram of total protein was observed in the cell extr acts of delta ALA-supplemented cells, The increase in heme biosynthesi s depended on the overexpression of a heme acceptor-CBS. Our data sugg est that bacterial heme synthesis is regulated beyond delta ALA syntha se. The delta ALA treatment resulted in 8 times more total CBS activit y with a 3.5-fold higher yield of the purified recombinant enzyme, mor e than 68% saturated with heme. Increased yield, higher specific activ ity, and improved heme saturation of CBS will facilitate large-scale p reparation of the enzyme, This method should be applicable to the over expression of other recombinant heme proteins in bacteria. (C) 1995 Ac ademic Press, Inc.