Purification, characterization, and genetic organization of recombinant Providencia stuartii urease expressed by Escherichia coli

Mulrooney, Scott B.; Lynch, Mary Jo; Mobley, Harry L. T.; Hausinger, Robert P.

doi:10.1128/jb.170.5.2202-2207.1988

Cited by 55 publications

(53 citation statements)

References 26 publications

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“…In regard to the restriction endonuclease cleavage sites determined, the three loci were indistinguishable. The maps presented here are in close agreement with the previously published restriction endonuclease map of the P. stuartii BE2467 urease gene cluster (22).…”

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confidence: 80%

Characterization of a plasmid-encoded urease gene cluster found in members of the family Enterobacteriaceae

D’Orazio

Collins

1993

J Bacteriol

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Plasmid-encoded urease gene clusters found in uropathogenic isolates of Escherichia coli, Providencia stuartii, and Salmonella cubana demonstrated DNA homology, similar positions of restriction endonuclease cleavage sites, and manners of urease expression and therefore represent the same locus. DNA sequence analysis indicated that the plasmid-encoded urease genes are closely related to the Proteus mirabilis urease genes.

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supporting

confidence: 80%

Characterization of a plasmid-encoded urease gene cluster found in members of the family Enterobacteriaceae

D’Orazio

Collins

1993

J Bacteriol

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“…Genetic analyses of the cloned ureases of Providencia stuartii and P. mirabilis have identified the coding regions for the structural subunits of the enzyme as well as the accessory polypeptides which are required for expression of enzyme activity in vivo (15,25,37). Mulrooney and co-workers (25) have purified the cloned urease of Providencia stuartii and determined its biochemical properties. In addition, they have demonstrated that the native enzyme possesses a heteromeric subunit structure of one large and two small polypeptides and contains four nickel ions per active enzyme molecule.…”

mentioning

confidence: 99%

“…1989, B64, p. 41) have been identified by cloning and expression in Escherichia coli. Genetic analyses of the cloned ureases of Providencia stuartii and P. mirabilis have identified the coding regions for the structural subunits of the enzyme as well as the accessory polypeptides which are required for expression of enzyme activity in vivo (15,25,37). Mulrooney and co-workers (25) have purified the cloned urease of Providencia stuartii and determined its biochemical properties.…”

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confidence: 99%

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Proteus mirabilis urease: nucleotide sequence determination and comparison with jack bean urease

1989

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Proteus mirabilis, a common cause of urinary tract infection, produces a potent urease that hydrolyzes urea to NH3 and C02, initiating kidney stone formation. Urease genes, which were localized to a 7.6-kilobase-pair region of DNA, were sequenced by using the dideoxy method. Six open reading frames were found within a region of 4,952 base pairs which were predicted to encode polypeptides of 31.0 (ureD), 11.0 (ureA), 12.2 (ureB), 61.0 (ureC), 17.9 (ureE), and 23.0 (ureF) kilodaltons (kDa). Each open reading frame was preceded by a ribosome-binding site, with the exception of ureE. Putative promoterlike sequences were identified upstream of ureD, ureA, and ureF. Possible termination sites were found downstream of ureD, ureC, and ureF. Structural subunits of the enzyme were encoded by ureA, ureB, and ureC and were translated from a single transcript in the order of 11.0, 12.2, and 61.0 kDa. When the deduced amino acid sequences of the P. mirabiis urease subunits were compared with the amino acid sequence of the jack bean urease, significant amino acid similarity was observed (58% exact matches; 73% exact plus conservative replacements). The 11.0-kDa polypeptide aligned with the N-terminal residues of the plant enzyme, the 12.2-kDa polypeptide lined up with internal residues, and the 61.0-kDa polypeptide matched with the C-terminal residues, suggesting an evolutionary relationship of the urease genes of jack bean and P. miabilis.Urinary tract infection with Proteus mirabilis can lead to serious complications, including cystitis, prostatitis, urolithiasis, pyelonephritis, bacteremia, and death (30, 38). The enzyme urease is recognized as an important virulence factor for this uropathogenic bacterial species and, indeed, as the causative agent of infection-induced kidney and bladder stones, which are estimated to represent 20 to 40% of all urinary stones (14). Alkalinization of the urine by hydrolysis of urea to carbon dioxide and ammonia facilitates precipitation of struvite, MgNH4PO4 6H20 and carbonateapatite, Ca1O(PO4CO3OH)6(OH). Furthermore, in catheterized patients, precipitation of urinary stones results in encrustation and blockage of indwelling urinary catheters. This complication has been uniquely correlated with the presence of P. mirabilis but not other ureolytic organisms (23). Further evidence suggests that the ammonia per se generated by ureolysis may be toxic to the kidney epithelia (5).Recent work has begun to yield an understanding of the biochemistry and genetics of ureases produced by members of the Proteeae tribe (21). Urease gene sequences from Providencia stuartii (22), P. mirabilis (15, 37), and Morganella morganii (L. Hu, B. Jones, M. Fox, E. Nicholson, and H. Mobley, Abstr. Annu. Meet. Am. Soc. Microbiol. 1989, B64, p. 41) have been identified by cloning and expression in Escherichia coli. Genetic analyses of the cloned ureases of Providencia stuartii and P. mirabilis have identified the coding regions for the structural subunits of the enzyme as well as the accessory polypeptides which are re...

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“…Urease genes have been isolated from Escherichia coli (7), Proteus mirabilis (20,41), Proteus vulgaris (30), Morganella morganni (15), Klebsiella pneumoniae (12), Klebsiella aerogenes (31), Providencia stuartii (29,32), Staphylococcus saprophyticus (11), Ureaplasma urealyticum (2,3), and Helicobacter pylori (5,24). In the members of the family Enterobacteriaceae, the genetic organization of the urease genes appears to be conserved in all species examined thus far.…”

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confidence: 99%

Insertional inactivation of an Escherichia coli urease gene by IS3411

Collins

Gutman

1992

J Bacteriol

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Ureolytic Escherichia coli are unusual clinical isolates that are found at various extraintestinal sites of infection, predominantly the urinary tract. The urease-positive phenotype is unstable in approximately 25% of these isolates, and urease-negative segregants are produced at a high frequency. We have studied the nature of the urease-positive-to-negative transition in one of these isolates, designated E. coli 1021. Southern hybridization experiments with genomic DNA extracted from seven independent E. coli 1021 urease-negative segregants revealed the presence of a 1.3-kb DNA insertion in the urease gene cluster. A DNA fragment containing the DNA insertion was cloned from one of the urease-negative segregants. This cloned DNA fragment was capable of mediating cointegrate formation with the conjugative plasmid pOX38, suggesting that the DNA insertion was a transposable element. The insert was identified as an IS3411 element in ureG by DNA sequence analysis. A 3-bp target duplication (CTG) flanking the insertion element was found. DNA spanning the insertion site was amplified from the other six urease-negative segregants by using the polymerase chain reaction. The DNA sequence of the amplified fragments indicated that an IS3411 element was found in an identical site in all urease-negative segregants examined. These data suggest that in E. coli 1021, IS3411 transposes at a high frequency into ureG at a CTG site, disrupting this gene and eliminating urease activity.

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Purification, characterization, and genetic organization of recombinant Providencia stuartii urease expressed by Escherichia coli

Cited by 55 publications

References 26 publications

Characterization of a plasmid-encoded urease gene cluster found in members of the family Enterobacteriaceae

Characterization of a plasmid-encoded urease gene cluster found in members of the family Enterobacteriaceae

Proteus mirabilis urease: nucleotide sequence determination and comparison with jack bean urease

Insertional inactivation of an Escherichia coli urease gene by IS3411

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