Literature summary for 3.5.1.24 extracted from

Xu, F.; Hu, X.J.; Singh, W.; Geng, W.; Tikhonova, I.G.; Lin, J.
The complex structure of bile salt hydrolase from Lactobacillus salivarius reveals the structural basis of substrate specificity (2019), Sci. Rep., 9, 12438 .

Cloned(Commentary)

Cloned (Comment)	Organism
gene bsh, recombinant expression of wild-type and mutant enzymes in Escherichia coli strain JL885	Ligilactobacillus salivarius

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified enzyme is crystallized from 16 mg/ml protein solution containing 10 mM sodium acetate, pH 5.5, 400 mM NaCl, 1 mM DTT, 1 mM EDTA, and 10% v/v glycerol by mixing with crystallization solution containing 20% w/v PEG 3350, 0.2 M KH2PO4, pH 4.8, at 20°C, crystals are soaked with substrate glycocholic acid for complex formation, X-ray diffraction structure determination and analysis at 2.10 A resolution, the complexed crystal contains enzyme with glycocholic acid and cholic acid, analysis of enzyme complex stability by molecular dynamics simulations	Ligilactobacillus salivarius

Crystallization (Comment)

Organism

purified enzyme is crystallized from 16 mg/ml protein solution containing 10 mM sodium acetate, pH 5.5, 400 mM NaCl, 1 mM DTT, 1 mM EDTA, and 10% v/v glycerol by mixing with crystallization solution containing 20% w/v PEG 3350, 0.2 M KH2PO4, pH 4.8, at 20°C, crystals are soaked with substrate glycocholic acid for complex formation, X-ray diffraction structure determination and analysis at 2.10 A resolution, the complexed crystal contains enzyme with glycocholic acid and cholic acid, analysis of enzyme complex stability by molecular dynamics simulations

Ligilactobacillus salivarius

Protein Variants

Protein Variants	Comment	Organism
C2S	site-directed mutagenesis, almost inactive mutant that shows very low remaining actiivty only with taurocholic acid	Ligilactobacillus salivarius
E270A	site-directed mutagenesis, the mutant shows increased activity and altered substrate specificity compared to wild-type	Ligilactobacillus salivarius
E270A	site-directed mutagenesis, the mutant shows reduced activity and altered substrate specificity compared to wild-type	Ligilactobacillus salivarius
F65A	site-directed mutagenesis, the mutant shows reduced activity and altered substrate specificity compared to wild-type	Ligilactobacillus salivarius
additional information	the pBSH plasmid bearing original bsh gene from Lactobacillus salivarius NRRL B-30514 is used as parent vector for site-directed mutagenesis. Construction of truncated mutant DELTA164-171, which is catalytically inactive	Ligilactobacillus salivarius
N171A	site-directed mutagenesis, the mutant shows reduced activity and altered substrate specificity compared to wild-type	Ligilactobacillus salivarius
Q257A	site-directed mutagenesis, the mutant shows reduced activity and altered substrate specificity compared to wild-type	Ligilactobacillus salivarius
Y24F	site-directed mutagenesis, the mutant shows reduced activity and altered substrate specificity compared to wild-type	Ligilactobacillus salivarius

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.
glycochenodeoxycholic acid + H2O	Ligilactobacillus salivarius	-	chenodeoxycholate + glycine	-	?
glycochenodeoxycholic acid + H2O	Ligilactobacillus salivarius NRRL B-30514	-	chenodeoxycholate + glycine	-	?
glycocholic acid + H2O	Ligilactobacillus salivarius	-	cholate + glycine	-	?
glycocholic acid + H2O	Ligilactobacillus salivarius NRRL B-30514	-	cholate + glycine	-	?
glycodeoxycholic acid + H2O	Ligilactobacillus salivarius	-	deoxycholate + glycine	-	?
glycodeoxycholic acid + H2O	Ligilactobacillus salivarius NRRL B-30514	-	deoxycholate + glycine	-	?
taurochenodeoxycholic acid + H2O	Ligilactobacillus salivarius	-	chenodeoxycholate + taurine	-	?
taurochenodeoxycholic acid + H2O	Ligilactobacillus salivarius NRRL B-30514	-	chenodeoxycholate + taurine	-	?
taurocholic acid + H2O	Ligilactobacillus salivarius	-	cholate + taurine	-	?
taurodeoxycholic acid + H2O	Ligilactobacillus salivarius	-	deoxycholate + taurine	-	?
taurodeoxycholic acid + H2O	Ligilactobacillus salivarius NRRL B-30514	-	deoxycholate + taurine	-	?

Organism

Organism	UniProt	Comment	Textmining
Ligilactobacillus salivarius	C7AQX8	-	-
Ligilactobacillus salivarius NRRL B-30514	C7AQX8	-	-

Posttranslational Modification

Posttranslational Modification	Comment	Organism
proteolytic modification	enzyme BSH undergoes an autocatalytic cleavage of the N-terminal residue to expose the cysteine residue to act as a nucleophile	Ligilactobacillus salivarius

Purification (Commentary)

Purification (Comment)	Organism
recombinant wild-type and mutant enzymes from Escherichia coli strain JL885	Ligilactobacillus salivarius

Source Tissue

Source Tissue	Comment	Organism	Textmining

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
glycochenodeoxycholic acid + H2O	-	Ligilactobacillus salivarius	chenodeoxycholate + glycine	-	?
glycochenodeoxycholic acid + H2O	-	Ligilactobacillus salivarius NRRL B-30514	chenodeoxycholate + glycine	-	?
glycocholic acid + H2O	-	Ligilactobacillus salivarius	cholate + glycine	-	?
glycocholic acid + H2O	-	Ligilactobacillus salivarius NRRL B-30514	cholate + glycine	-	?
glycodeoxycholic acid + H2O	-	Ligilactobacillus salivarius	deoxycholate + glycine	-	?
glycodeoxycholic acid + H2O	-	Ligilactobacillus salivarius NRRL B-30514	deoxycholate + glycine	-	?
taurochenodeoxycholic acid + H2O	-	Ligilactobacillus salivarius	chenodeoxycholate + taurine	-	?
taurochenodeoxycholic acid + H2O	-	Ligilactobacillus salivarius NRRL B-30514	chenodeoxycholate + taurine	-	?
taurocholic acid + H2O	-	Ligilactobacillus salivarius	cholate + taurine	-	?
taurodeoxycholic acid + H2O	-	Ligilactobacillus salivarius	deoxycholate + taurine	-	?
taurodeoxycholic acid + H2O	-	Ligilactobacillus salivarius NRRL B-30514	deoxycholate + taurine	-	?

Subunits

Subunits	Comment	Organism
More	overall structures of GCA-soaked lsBSH complex. It consists of two tetramers, which are composed of chains A, B, C, D, E, F, G and H. In each lsBSH tetramer, one monomer is in complex with the substrate glycocholic acid, while the remaining monomers are in complex with the product cholic acid. Molecular dynamic simulations using lsBSH-GCA complex (PDB ID 5Y7P) as template. Structure comparisons, overview	Ligilactobacillus salivarius

Synonyms

Synonyms	Comment	Organism
bile salt hydrolase	-	Ligilactobacillus salivarius
BSH	-	Ligilactobacillus salivarius
lsBSH	-	Ligilactobacillus salivarius

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
37	-	assay at	Ligilactobacillus salivarius

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
6	-	assay at	Ligilactobacillus salivarius

General Information

General Information	Comment	Organism
additional information	the active site in lsBSH is located in a shallow and water-exposed cavity formed by beta-sheets and four loops, loops 1-4. Loops 2 and 3 shape and partially close the active site of lsBSH from water exposure. Cys2 and Asn171 are critical for enzymatic activity, while Tyr24, Phe65 and Gln257 contribute to the substrate specificity. Structural insights into BSH-substrate interactions, the mechanism of catalysis, and substrate specificity, overview. Enzyme BSH is an N-terminal nucleophilic (Ntn) hydrolase. The enzyme undergoes an autocatalytic cleavage of the N-terminal residue to expose the cysteine residue to act as a nucleophile	Ligilactobacillus salivarius