DETERMINATION OF SERUM BIOTINIDASE ACTIVITY WITH BIOTINYL DERIVATIVESOF IODOTYRAMINES AS SUBSTRATES

We synthesized biotinylated mono- and di-iodotyramine and their radioa ctive counterparts and used these substances as substrates to estimate serum biotinidase activity in a radioassay system. The K-m values det ermined for mono- and di-iodobiotinyl derivatives were 15.8 and 25.9 m u M, respectively, whereas, the maximum velocities of the enzymatic re action were 27.0 and 8.7 nmol . min(-1) . mL(-1), respectively. Both s ubstrates competed with biocytin for the same active site of the enzym e and the K-l values were 7.30 and 9.56 mu M for the mono- and di-iodi nated substrate, respectively. Higher assay sensitivity was obtained u sing [I-125]biotinyl-monoiodotyramine as substrate, and the values obt ained were directly related with those determined with the well-establ ished colorimetric method (r = 0.9377, n = 31). However, for routine u se, the assay may be accomplished by diluting the radiotracer with bio cytin instead of its ''cold'' counterpart, because it is a commerciall y available reagent. The values obtained in this case were very well c orrelated with those determined by the colorimetric assay as well (r = 0.9289, n = 31).