COMPARATIVE-STUDIES OF HUMAN RECOMBINANT 74- AND 54-KDA L-HISTIDINE DECARBOXYLASES

We have expressed and characterized human recombinant 74-kDa (rHDC74) and 54-kDa (rHDC54) L-histidine decarboxylases (HDCs) in Sf9 cells, By immunoblot analysis, rHDC74 and rHDC54 were shown to be localized pre dominantly in the particulate and soluble fractions, respectively, rHD C74 exhibited histamine-synthesizing activity equivalent to that of rH DC54. The existence of 74- and 54-kDa HDCs was also confirmed in the p articulate and supernatant fractions of the cell lysate, respectively, from the human basophilic leukemia cell line KU-812-F. The ratio of H DC activity to immunoreactivity was similar for the two forms of the e nzyme. The specific activity of purified rHDC54 (1.12 mu mol/mg/min) w as comparable to those of HDCs from other mammalian tissues or cells. The purified rHDC54 was eluted as a monomer form from a Superdex-200 c olumn; the molecular mass of the enzyme was approximately 54 kDa on SD S-polyacrylamide gel electrophoresis without 2-mercaptoethanol. The HD C activity of rHDC54 significantly decreased on dialysis against buffe r without pyridoxal 5'-phosphate; addition of pyridoxal 5'-phosphate t o the dialysate readily increased in the enzyme activity to the origin al activity. Taken together, these results suggest that human HDC func tions as both 74- and 54-kDa forms having equivalent HDC activity, whi ch are localized in the particulate and soluble fractions, respectivel y, and that the latter form exhibits its activity as a monomer form.