Comparative study of human milk and serum biotinidase

Abstract Biotinidase was purified from human breast milk (4 000-fold), and was compared with human serum biotinidase (enriched 30 000-fold). The molecular weight of milk enzyme was 68 000 Da is determined by SDS-PAGE. It was definitely smaller than that of serum biotinidase ( M r = 76 000). Isoelectric point of milk biotinidase was 4.6, whereas that of serum biotinidase was 4.3. Sialic acid pontent in milk biotinidase was less than that found in serum enzyme. N -Acetyl-galactosamine was present in milk enzyme, whereas it was absent in serum enzyme. Milk biotinidase is O -glycosylated, thereas serum biotinidase is N -glycosylated. These differences in glycosylation suggest the existence of different types of excretion mechanisms between milk and serum biotinidase. Both biotinidases were found to be thiol-type enzyme, however, the extent of activation of the enzemy 2-mercaptoethanol was 13-fold in milk, whilt the serum enzyme was activated only 1.5-fold. K m for biotinyl-4-amino-benzoate was 22 μM in milk enzyme and 50 μM in seruum enzyme. Competitive inhibition by biotin ( K i ) of milk enzyme was 43 μM and 1.3 mM for serum enzyme. These results suggeest the structural differences it or near the active site of the each enzyme.