Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.5.1.12 (biotinidase)
 392 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Late-onset multiple carboxylase deficiency depends on biotinidase deficiency and is inherited as an autosomal recessive trait. Lipoamidase deficiency in humans has not been previously reported, using the natural substrate lipoyllysine for lipoamidase. In this report we describe a simultaneous decrease in both lipoamidase and biotinidase activity in serum from a 21 month-old boy with a profound biotinidase deficiency. Lipoamidase activity in human serum was determined with both lipoyllysine (epsilon-N-(D,L-lipoyl)-L-lysine) and N-D,L-lipoyl-p-aminobenzoate as substrates. Biotinidase activity was determined with both biocytin (epsilon-N-(D-biotinyl-L-lysine) and N-D-biotinyl-p-aminobenzoate as substrates. Our findings indicate that lipoamidase activity and biotinidase activity in human serim are due to the same enzyme, but a "residual activity" was usually found when N-D,L-lipoyl-p-aminobenzoate was used as a substrate. Compared with the activity in control serum, this "residual activity" was little affected by inhibition with biocytin, indicating that a small fraction of a modified biotinidase probably exists.
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PMID:Lipoamidase and biotinidase deficiency: evidence that lipoamidase and biotinidase are the same enzyme in human serum. 159 76

A radioassay for determining biotinidase activity in human serum was developed, using N-[beta-(4-OH-3-125I-phenyl)ethyl]-biotinamide in combination with biocytin as the substrate, avidin as a binding protein, and polyethylene glycol as a separation reagent. The gamma-emitting 125I-biotinamide (= tracer) was synthesized by coupling (pH 8.5, 20-22 degrees C, 90 min) N-hydroxysuccinimidobiotin to 125I-tyramine. Using polyethylene glycol as a separation reagent, it was possible to eliminate several problems that were encountered when other separation reagents were used. Biotinidase activity was evaluated following the cleavage of the 125I-biotinamide and expressed in fmol of tracer cleaved.min-1.ml-1 in the presence of 9 nmol of biocytin. Under the conditions used, the time response of the assay was linear up to 3 h. The method is simple to perform, more sensitive than the previously described methods, and reproducible (intra- and interassay CVs of 4.9 and 10.2%, respectively) and allows the simultaneous handling of more than 100 samples in less than 3 h.
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PMID:Biotinidase radioassay using an 125I-biotin derivative, avidin, and polyethylene glycol reagents. 177 89

Biotinidase activity was measured in plasmas of 1-, 7-, 14-, and 21-day-old rats from control dams and dams that had been fed a biotin-depleting diet from Day 15 of gestation. Biotinidase activity increased significantly in the plasma of rats from control and depleted mothers until Postnatal Day 14, after which there was a small but significant decline at Day 21. Differences between the mean activities of the two groups of pups on each sampling day were not significant and there were no significant differences in activity levels attributable to sex. Plasma albumin concentrations increased from birth until Day 21, and plasma biotinidase activity and albumin concentration were significantly correlated (r = +/- 0.43). We suggested that these two proteins may be controlled by a common mechanism in the early postnatal period, and that biotin deficiency does not affect the development of biotinidase activity. Because biotin-depleted neonatal pups show developmental changes in biotinidase activity similar to those of human newborns, and they can be produced reliably by depleting dams from Day 15 of gestation, they may be useful models for studying the developmental abnormalities associated with human biotinidase deficiency.
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PMID:Effects of age and biotin status on postnatal development of plasma biotinidase activity in rats. 201 13

Biotinidase activities found in porcine brains (n = 3) were as follows: cerebrum, 4.4 +/- 0.2 pmol/min per milligram of protein; cerebellum, 7.6 +/- 0.3 pmol/min per milligram of protein; medulla, 2.9 +/- 0.3 pmol/min per milligram of protein. These values are relatively high compared with the activities in rat or guinea pig brains. Subcellular distribution of biotinidase was found mainly in the soluble cytoplasmic fraction (S3), i.e., in the supernatant of 0.32 M sucrose S2 solution after ultracentrifugation at 105,000g for 90 min. This is in contrast to the guinea pig livers, in which the subcellular distribution of biotinidase is mainly found in the microsomal fraction. After a seven-step purification (22,200-fold enrichment), porcine brain biotinidase is identified as a single polypeptide by the sodium dodecyl sulfate-polyacrylamide gel electrophoresis system, and its molecular weight is determined as 68,000 Da. The isoelectric point of the enzyme was 4.3. Sialidase treatment strongly suggests the presence of sialyl residues in this enzyme. Amino acid analysis indicates relatively high hydrophilicity and high content of glycine and serine. The enzyme activity is inhibited by organic mercurials, but not by diisopropylfluorophosphate. Abundant soluble biotinidase in brain cytoplasm may play an important role which has not been discovered yet.
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PMID:Biotinidase in the porcine cerebrum. 232 94

Biotinidase (EC 3.5.1.12) activity in human breast milk was first demonstrated by us, using a newly developed method. The enzyme activity was relatively low compared with that in human serum. However, its activity increased significantly with thiol-reactive agents. Taurine and glutathione also enhanced the enzyme activity considerably. From these results, we speculate that biotinidase in milk would be protected from various inactivation-mechanisms in the same manner as milk lipase. Biotinidase might play an important role in the intestinal absorption of the indispensable vitamin, biotin, during infancy, when enzymes in the pancreatic juice are still immature.
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PMID:The significance of breast milk biotinidase. 251 65

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 as determined by SDS-PAGE. It was definitely smaller than that of serum biotinidase (Mr = 76,000). Isoelectric point of milk biotinidase was 4.6, whereas that of serum biotinidase was 4.3. Sialic acid content 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, whereas 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 enzyme by 2-mercaptoethanol was 13-fold in milk, whilst the serum enzyme was activated only 1.5-fold. Km for biotinyl-4-amino-benzoate was 22 microM in milk enzyme and 50 microM in serum enzyme. Competitive inhibition by biotin (Ki) of milk enzyme was 43 microM and 1.3 mM for serum enzyme. These results suggest the structural differences at or near the active site of the each enzyme.
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PMID:Comparative study on human milk and serum biotinidase. 251 77

Thirty-two human serum specimens were assayed for lipoamidase (lipoyl-4-aminobenzoate hydrolase) activity. All sera had lipoamidase activities. This substrate was newly synthesized by us and had a satisfying purity as evaluated by HPLC-fluorimetric detection. Product (p-aminobenzoate) liberated was determined directly by the HPLC-fluorimetric method. Liberation of the product was linearly continued for 6 h. The pH optimum of serum lipoamidase was found to be 7.0. The effect of substrate concentration on human serum lipoamidase activity was examined and the reaction was saturated at 0.1 mmol/l. The sera obtained were from individuals aged from 1 to 8 years. The mean value of serum lipoamidase activity was found to be 1.50 U/l (SD 1.037, range 0.04-3.75, n = 32). The difference of sex effects was analyzed and no significant difference was found (males: n = 14, mean 1.48, SD 1.162, range 0.04-3.75; females: n = 18, mean 1.52, SD 0.963, range 0.48-3.51) among this age group. Biotinidase activity was also determined in these 32 serum specimens and the correlation was examined. The mean biotinidase activity was 3.16 U/l (SD 2.567, range 0.35-9.37). The correlation coefficient (r) between lipoamidase activity and biotinidase activity was 0.8931. Although the physiological significance of lipoamidase has not been known, the enzyme might play an important role in recycling of lipoate as biotinidase does.
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PMID:Human serum lipoamidase. 316 70

The effects of a disulfide reducing agent and sulfhydryl blocking agents on the biotinidase activity in human serum and on the purified biotinidase have been extensively studied by using a newly developed HPLC assay method. This HPLC method directly measures the product (p-aminobenzoate, PAB), and is not interfered with by sulfhydryl-reactive agents. Further, because the substrate solution of this HPLC assay method contains only substrate (biotin 4-amidobenzoate) and phosphate buffer, accurate studies on the effects of sulfhydryl blocking reagents on the purified enzyme could be performed. Biotinidase activities in human sera (n = 83) were always enhanced by 2-mercaptoethanol (ME). The optimum concentration was found to be 1 mM. The degree of activation was variable (100 to 400% of the original) depending on the serum sample. Sulfhydryl blocking reagents such as organic mercurials were tested on fresh serum and purified enzyme. Mercuric agents were found to inhibit the activity of fresh serum and purified enzyme at 0.05 and 0.005 mM, respectively. Sulfhydryl alkylating agents, N-ethylmaleimide (NEM) and dithiobis(2-nitro)benzoic acid (DTNB), inhibited 100 and 64% of the activity of the purified enzyme at 0.1 and 1.0 mM, respectively. However, lower concentrations (less than 5 nM) of organic mercurials and mercuric ion exhibited a slight enhancement (20-30%) of the activity of the purified enzyme. These results indicate the presence of an essential sulfhydryl residue at the active center. The enzyme contains 2.5 sulfhydryls per molecule, as determined by using Ellman's assay method. Serine protease inhibitors such as phenylmethylsulfonyl fluoride (PMSF) and diisopropylfluorophosphate (DFP) did not inhibit the enzyme activity at 0.05 mM or higher concentration.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Human serum biotinidase is a thiol-type enzyme. 318 46

Biotinidase shows two binding sites for biotin, with Kd = 59 and 3 nM respectively, and requires tryptophan and cysteine residues of the biotinidase protein for biotin-binding activity. Analysis of human serum by various column-chromatographic techniques indicates that biotinidase is the only protein which exchanges with labelled (+)-biotin. It was shown previously that epileptic patients receiving a high average dose of anticonvulsants (containing a carbamide group) have lower biotin concentrations than those receiving a low dose. We have shown in human serum and with purified biotinidase that these anticonvulsant drugs compete with biotin for binding to the protein moiety.
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PMID:Role of human serum biotinidase as biotin-binding protein. 322 2

Biotinidase activity in 19 samples of human breast milk was investigated with the sensitive high-performance liquid chromatography (HPLC)-fluorometric method that we developed. All samples exhibited biotinidase activity. For mature milk the mean activity of 17 samples was 0.208 nmol.min-1.mL-1 milk (range, 0.087-0.516 nmol.min-1.mL-1) and mean specific activity was 7.51 pmol.min-1.mg-1 protein (range, 2.17-17.2 pmol.min-1.mg-1). These values are relatively low compared with the activity in human serum (5.26 +/- 2.92 nmol.min-1.mL-1 serum and 95.6 +/- 53.1 pmol.min-1.mg-1 protein; n = 246). Biotinidase activities of milk obtained at various times after birth were not significantly different. However, biotinidase activity in colostrum was about five times higher than that of mature milk. The existence of biotinidase activity in all specimens suggests that this enzyme plays an important nutritional role during infancy.
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PMID:Biotinidase in human breast milk. 340 8


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