Kinetic properties of 7α-hydroxysteroid dehydrogenase from Escherichia coli 080

The effect of Escherichia coli K-12 on the transformation of hyocholic acid, α-muricholic acid, β-muricholic acid and 3α,7β,12α-trihydroxy-5β-cholanoic acid was examined in aerobic and anaerobic cultures for 4 days and the activities were compared with those on cholic acid and chenodeoxycholic acid. Hyocholic acid was dehydrogenated, but to a lesser extent than cholic acid or chenodeoxycholic acid. However, α-muricholic acid, which has a 7α-hydroxyl group, was not dehydrogenated even after 4 days. This was confirmed by experiments using a cell-free crude enzyme preparation and by gas chromatographic analysis of bile acids in the medium culture. β-Muricholic acid and 3α,7β,12α-trihydroxy-5β-cholanoic acid were not changed by Esherichia coli K-12.

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“…It is reported that 7α-HSDH has a higher affinity for CDCA than for CA (6,12), and this was confirmed in our experiments (Fig. 2).…”

Section: Discussionsupporting

confidence: 91%

Aerobic and Anaerobic Biotransformation of Bile Acids by Escherichia coli (II): No Conversion of α-Muricholic Acid by 7α-Hydroxysteroid Dehydrogenase of E. coli

Ogura

Yamaga

Suzuki

et al. 2005

Bioscience Microflora

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“…7a-HSDHs generally use NADP(H) as a cofactor, with the exception of E. coli (85) and Ba. thetaiotaomicron (74).…”

Section: Microbial Bile Acid Hydroxysteroid Dehydrogenase(s)mentioning

confidence: 99%

“…Regulation of 7a/b-HSDH expression is generally growth phase-dependent and inducible by bile acid substrates (Table 2). C. scindens and E. coli constitutively express 7a-HSDHs and are noninducible (69,85). Unexpectedly, the nonsubstrate DCA can induce 7a-HSDH expression in C. absonum and C. sordellii, although the reason for this induction remains unclear (70,76).…”

Section: Microbial Bile Acid Hydroxysteroid Dehydrogenase(s)mentioning

confidence: 99%

Bile salt biotransformations by human intestinal bacteria

Ridlon

Kang

Hylemon

2006

Journal of Lipid Research

2,198

2,417

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Secondary bile acids, produced solely by intestinal bacteria, can accumulate to high levels in the enterohepatic circulation of some individuals and may contribute to the pathogenesis of colon cancer, gallstones, and other gastrointestinal (GI) diseases. Bile salt hydrolysis and hydroxy group dehydrogenation reactions are carried out by a broad spectrum of intestinal anaerobic bacteria, whereas bile acid 7-dehydroxylation appears restricted to a limited number of intestinal anaerobes representing a small fraction of the total colonic flora. Microbial enzymes modifying bile salts differ between species with respect to pH optima, enzyme kinetics, substrate specificity, cellular location, and possibly physiological function. Crystallization, site-directed mutagenesis, and comparisons of protein secondary structure have provided insight into the mechanisms of several bile acid-biotransforming enzymatic reactions. Molecular cloning of genes encoding bile salt-modifying enzymes has facilitated the understanding of the genetic organization of these pathways and is a means of developing probes for the detection of bile salt-modifying bacteria. The potential exists for altering the bile acid pool by targeting key enzymes in the 7a/b-dehydroxylation pathway through the development of pharmaceuticals or sequestering bile acids biologically in probiotic bacteria, which may result in their effective removal from the host after excretion.-Ridlon, J. M., D-J. Kang, and P. B. Hylemon. Bile salt biotransformations by human intestinal bacteria. J. Lipid Res. 2006. 47: 241-259.

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“…7a-Hydroxysteroid dehydrogenases have been purified from Bacteroides species (23,29,41), Escherichia coli (28,36,37), and rat liver microsomes (1). Recently, two 7a-hydroxysteroid dehydrogenases have been purified to homogeneity and extensively characterized: an NAD-linked enzyme from E. coli HB101, which has been cloned, sequenced, and * Corresponding author.…”

mentioning

confidence: 99%

Cloning, sequencing, and expression of the gene coding for bile acid 7 alpha-hydroxysteroid dehydrogenase from Eubacterium sp. strain VPI 12708

1991

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Southern blot analysis indicated that the gene encoding the constitutive, NADP-linked bile acid 7a-hydroxysteroid dehydrogenase of Eubacterium sp. strain VPI 12708 was located on a 6.5-kb EcoRI fragment of the chromosomal DNA. This fragment was cloned into bacteriophage lambda gtll, and a 2.9-kb piece of this insert was subcloned into pUC19, yielding the recombinant plasmid pBH51. DNA sequence analysis of the 7a-hydroxysteroid dehydrogenase gene in pBH51 revealed a 798-bp open reading frame, coding for a protein with a calculated molecular weight of 28,500. A putative promoter sequence and ribosome binding site were identified. The 7a-hydroxysteroid dehydrogenase mRNA transcript in Eubacterium sp. strain VPI 12708 was about 0.94 kb in length, suggesting that it is monocistronic. An Escherichia coli DH5a transformant harboring pBH51 had approximately 30-fold greater levels of 7a-hydroxysteroid dehydrogenase mRNA, immunoreactive protein, and specific activity than Eubacterium sp. strain VPI 12708. The 7af-hydroxysteroid dehydrogenase purified from the pBH51 transformant was similar in subunit molecular weight, specific activity, and kinetic properties to that from Eubacterium sp. strain VPI 12708, and it reacted with antiserum raised against the authentic enzyme on Western immunoblots. Alignment of the amino acid sequence of the 7a-hydroxysteroid dehydrogenase with those of 10 other pyridine nucleotide-linked alcohol/polyol dehydrogenases revealed six conserved amino acid residues in the N-terminal regions thought to function in coenzyme binding.Many members of the human intestinal microflora can modify the glycine and taurine conjugates of the primary bile acids, cholic acid and chenodeoxycholic acid, producing up to 20 different bile acid metabolites (21). The predominant modifications include deconjugation of the glycine and taurine moieties by bile salt hydrolase, removal of 7-hydroxy groups (7-dehydroxylation), and dehydrogenation of a-and ,B-hydroxy groups at carbons 3, 7, and 12 by stereospecific bile acid hydroxysteroid dehydrogenases to yield oxo bile acids.Quantitatively, the most important of these transformations is the 7ux-dehydroxylation of cholic and chenodeoxycholic acids, producing deoxycholic and lithocholic acids, respectively (43). The human intestinal anaerobic bacterium, Eubacterium sp. strain VPI 12708, has a multistep 7a-dehydroxylation pathway which is induced by addition of cholic acid to the culture medium (6,9,11, 22,(46)(47)(48). Several bile acid-inducible (bai) genes encoding proteins involved in this pathway have been cloned and sequenced (10,11,18,31,49,50).7a-Hydroxysteroid dehydrogenase is the most common class of bile acid hydroxysteroid dehydrogenases found in nature (3), having been detected in numerous genera of bacteria (3, 14-16, 28, 30, 37) and in mammalian liver (1). 7a-Hydroxysteroid dehydrogenases have been purified from Bacteroides species (23,29,41), Escherichia coli (28,36,37), and rat liver microsomes (1). Recently, two 7a-hydroxysteroid dehydrogenases have been pu...

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Kinetic properties of 7α-hydroxysteroid dehydrogenase from Escherichia coli 080

Cited by 16 publications

References 6 publications

Aerobic and Anaerobic Biotransformation of Bile Acids by <i>Escherichia coli</i> (II): No Conversion of α-Muricholic Acid by 7α-Hydroxysteroid Dehydrogenase of <i>E. coli</i>

Aerobic and Anaerobic Biotransformation of Bile Acids by <i>Escherichia coli</i> (II): No Conversion of α-Muricholic Acid by 7α-Hydroxysteroid Dehydrogenase of <i>E. coli</i>

Bile salt biotransformations by human intestinal bacteria

Cloning, sequencing, and expression of the gene coding for bile acid 7 alpha-hydroxysteroid dehydrogenase from Eubacterium sp. strain VPI 12708

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