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

Recently, Hymes and co-workers demonstrated that human biotinidase (EC 3.5.1.12) specifically biotinylates histones, suggesting that biotin may have a specific role in transcription and replication of DNA. In the present study, we sought to biotinylate histones in vitro for later use as standards in the quantitation of histones biotinylated in vivo. We also sought to develop a procedure for electrophoretic separation and streptavidin-peroxidase detection of the various classes of biotinylated histones. Histones H1, H2a, H2b, H3, and H4 from calf thymus were biotinylated using sulfosuccinimidobiotin at pH 7.5. Stoichiometries of biotin/histone were determined either by 4'-hydroxyazobenzene-2-carboxylic acid/avidin assay or by avidin-binding assay. The stoichiometries of biotinylation (mol biotin/mol histone) were as follows: H1, 3.9 +/- 0.17; H2a, 1.7 +/- 0.11; H2b, 1.8 +/- 0.11; H3, 0.029 +/- 0.0012; H4, 0.006 +/- 0.0002. When two synthetic polypeptides were used as substrates for biotinylation, the stoichiometry of poly-l-lysine was 2.8 +/- 0.14 mol biotin/mol; in contrast, the stoichiometry of poly-l-arginine was less than 0.3 x 10(-3) mol biotin/mol. These data suggest that primary amino groups of histones biotinylated by sulfosuccinimidobiotin were lysine rather than arginine. Detection and identification of biotinylated histones were accomplished by electrophoretic separation on 16% polyacrylamide gels; the separated histones on nitrocellulose transblots of the gels were detected using streptavidin-peroxidase with 4-chloro-1-naphthol as the substrate. We conclude that sulfosuccinimidobiotin does biotinylate each of the five classes of histones and that the stoichiometry of biotinylation is sufficient for detection on nitrocellulose transblots by streptavidin-peroxidase.
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PMID:Chemical synthesis of biotinylated histones and analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis/streptavidin-peroxidase. 1052 92

Folding of DNA into chromatin is mediated by binding to histones such as H4; association of DNA with histones is regulated by covalent histone modifications, e.g. acetylation, methylation, and biotinylation. We sought to identify amino-acid residues that are biotinylated in histone H4, and to determine whether acetylation and methylation of histones affect biotinylation. Synthetic peptides spanning fragments of human histone H4 were biotinylated enzymatically using biotinidase. Peptide-bound biotin was probed with streptavidin-peroxidase. Peptides based on the N-terminal sequence of histone H4 were effectively recognized by biotinidase as substrates for biotinylation; in contrast, peptides based on the C-terminal sequences were not biotinylated. Substitution of K8 or K12 with alanine or arginine decreased biotinylation, suggesting that these lysines are targets for biotinylation; K8 and K12 are also known targets for acetylation. Chemical acetylation or methylation of a given lysine decreased subsequent enzymatic biotinylation of neighboring lysines, consistent with cross-talk among histone modifications. Substitution of a given lysine (positive charge) with glutamate (negative charge) abolished biotinylation of neighboring lysines, providing evidence that the net charge of histones has a role in biotinylation. An antibody was generated that specifically recognized histone H4 biotinylated at K12. This antibody was used to detect biotinylated histone H4 in nuclear extracts from human cells. These studies suggest that K8 and K12 in histone H4 are targets for biotinylation, that acetylation and biotinylation compete for the same binding sites, and that acetylation and methylation of histones affect biotinylation of neighboring lysines.
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PMID:K8 and K12 are biotinylated in human histone H4. 1515 16

Histones are modified post-translationally, e.g. by methylation of lysine and arginine residues, and by phosphorylation of serine residues. These modifications regulate processes such as gene expression, DNA repair, and mitosis and meiosis. Recently, evidence has been provided that histones are also modified by covalent binding of the vitamin biotin. The aims of this study were to identify biotinylation sites in histone H3, and to investigate the crosstalk among histone biotinylation, methylation and phosphorylation. Synthetic peptides based on the sequence of human histone H3 were used as substrates for enzymatic biotinylation by biotinidase; biotin in peptides was probed using streptavidin peroxidase. These studies provided evidence that K4, K9 and K18 in histone H3 are good targets for biotinylation; K14 and K23 are relatively poor targets. Antibodies were generated to histone H3, biotinylated either at K4, K9 or K18. These antibodies localized to nuclei in human placental cells in immunocytochemistry and immunoblotting experiments, suggesting that lysines in histone H3 are biotinylated in vivo. Dimethylation of R2, R8 and R17 increased biotinylation of K4, K9 and K18, respectively, by biotinidase; phosphorylation of S10 abolished biotinylation of K9. These observations are consistent with crosstalk between biotinylation of histones and other known modifications of histones. We speculate that this crosstalk provides a link to known roles for biotin in gene expression and cell proliferation.
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PMID:K4, K9 and K18 in human histone H3 are targets for biotinylation by biotinidase. 1609 5