title: Molecular cloning, characterization and comparison of bile salt hydrolases from Lactobacillus johnsonii PF01.
abstract: OBJECTIVE
  To clone, characterize and compare the bile salt hydrolase (BSH) genes of Lactobacillus johnsonii PF01.
  
  
  RESULTS
  The BSH genes were amplified by polymerase chain reaction (PCR) using specific oligonucleotide primers, and the products were inserted into the pET21b expression vector. Escherichia coli BLR (DE3) cells were transformed with pET21b vectors containing the BSH genes and induced using 0·1 mmol l(-1)  isopropylthiolgalactopyranoside. The overexpressed BSH enzymes were purified using a nickel-nitrilotriacetic acid (Ni(2+) -NTA) agarose column and their activities characterized. BSH A hydrolysed tauro-conjugated bile salts optimally at pH 5·0 and 55°C, whereas BSH C hydrolysed glyco-conjugated bile salts optimally at pH 5·0 and 70°C. The enzymes had no preferential activities towards a specific cholyl moiety.
  
  
  CONCLUSIONS
  BSH enzymes vary in their substrate specificities and characteristics to broaden its activity. Despite the lack of conservation in their putative substrate-binding sites, these remain functional through motif conservation.
  
  
  CONCLUSIONS
  This is to our knowledge the first report of isolation of BSH enzymes from a single strain, showing hydrolase activity towards either glyco-conjugated or tauro-conjugated bile salts. Future structural homology studies and site-directed mutagenesis of sites associated with substrate specificity may elucidate specificities of BSH enzymes.
authors.affiliation: Department of Animal Resources Science, Dankook University, Cheonan, Korea.
authors.forename: J P
authors.initials: JP
authors.lastname: Chae
authors.forename: V D
authors.initials: VD
authors.lastname: Valeriano
authors.forename: G-B
authors.initials: GB
authors.lastname: Kim
authors.forename: D-K
authors.initials: DK
authors.lastname: Kang
mesh_terms: D000581:Amidohydrolases / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D001483:Base Sequence; D001647:Bile Acids and Salts / Q000378:metabolism*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005798:Genes, Bacterial; D006863:Hydrogen-Ion Concentration; D007778:Lactobacillus / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D001647:Bile Acids and Salts; D000581:Amidohydrolases; C023154:choloylglycine hydrolase