Penicillin acylase from Alcaligenes faecalis has a very high affinity for both natural (benzylpenicillin, K m -0.0042 mM) and colorimetric (6-nitro-3-phenylacetamidobenzoic acid, K m = 0.0045 mM) substrates as well as the product of their hydrolysis, phenylacetic acid (A\ = 0.016 mM). The enzyme is partially inhibited at high benzylpenicillin concentrations but the triple SES complex formed still retains 43% of the maximal catalytic activity; the affinity of benzylpenicillin for the second substrate molecule binding site is much lower (As' = 54 mM) than for the first one. Phenylmethylsulfonyl fluoride was shown to be a very effective irreversible inhibitor, completely inactivating the penicillin acylase from A. faecalis in a few minutes at micromolar concentrations; this compound was used for enzyme active site titration. The absolute values of the determined kinetic parameters for enzymatic hydrolysis of 6-nitro-3-phenylacetamidobenzoic acid (k cat = 95 s _1 and k cat IK m = 2.1 X10 7 M _1 s _1 ) and benzylpenicillin (A cat = 54 s _1 and k cat IK m -1.3 X10 -7 M 1 s 1 ) by penicillin acylase from A. faecalis were shown to be highest of all the enzymes of this family that have so far been studied.

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Both favorable pregnancy outcomes and fetal abnormalities have been associated with the use of tyrosine kinase inhibitors (TKIs) during pregnancy. The placental transfer of TKIs in humans is poorly understood. We observed women with chronic myeloid leukemia who used imatinib or nilotinib during the late pregnancy stages. The newborns had no birth abnormalities. We evaluated the drug concentrations in maternal blood, umbilical cord blood, and placental samples collected during labor. We found limited placental transfer of the TKIs. The fetal/maternal concentration ratio ranged from 0.5 to 0.58 for nilotinib and from 0.05 to 0.22 for imatinib. The placental/maternal ratio was higher for imatinib than for nilotinib. Theoretical pharmacokinetic modeling of passive placental crossing was insufficient to predict the in vivo data because the calculated fetal/maternal ratio was close to 1 for both drugs. We propose that active placental transport contributes to fetal protection against TKI exposure during pregnancy.

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