In Vivo and in Vitro Kinetics of Metal Transfer by the Klebsiella aerogenes Urease Nickel Metallochaperone, UreE

Colpas, Gerard J.; Hausinger, Robert P.

doi:10.1074/jbc.275.15.10731

Cited by 60 publications

(66 citation statements)

References 35 publications

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“…The UreE dimer binds three copper ions (Fig. 1a), consistent with the reported three sequential Cu 2ϩ binding steps observed by kinetics (18). The pair of His-96 residues binds one copper atom between the subunits, whereas the other two copper sites involve His-110 and His-112 from within each subunit (Fig.…”

Section: Resultssupporting

confidence: 75%

“…These results parallel the reported inability to crystallize full-length wild-type UreE with nickel and the fracturing of wild-type apoprotein crystals upon nickel addition (14). As an alternative to obtaining the nickel-bound structure, we solved the copperbound form of H144*UreE for which the metal binding properties are well characterized (17,18). After soaking 50 mM CuSO 4 solution into pre-grown UreE crystals for 2 h, the copperbinding sites were readily located in the (F o ϪF c ) difference Fourier and anomalous difference maps (Fig.…”

Section: Resultssupporting

confidence: 59%

“…2, a and b). While the Atx1 thiol is one of two cysteines essential to its copper metallochaperone role, Cys-79 is not important to function of UreE (17,18). Cys-89, located in helix H2, has no counterpart in Atx1 and is not available to bind metals because it is involved in subunit dimerization.…”

Section: Resultsmentioning

confidence: 99%

“…The metal-binding properties of wild-type UreE and H144*UreE, which has 15 residues truncated C-terminally, have been extensively characterized (15)(16)(17)(18). The wild-type protein binds approximately six nickel ions per homodimer in distorted octahedral geometry with an average of three-five histidine donors per metal ion (14,16).…”

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confidence: 99%

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Crystal Structure of Klebsiella aerogenesUreE, a Nickel-binding Metallochaperone for Urease Activation

Song

Mulrooney

Huber

et al. 2001

Journal of Biological Chemistry

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UreE is proposed to be a metallochaperone that delivers nickel ions to urease during activation of this bacterial virulence factor. Wild-type Klebsiella aerogenes UreE binds approximately six nickel ions per homodimer, whereas H144*UreE (a functional C-terminal truncated variant) was previously reported to bind two. We determined the structure of H144*UreE by multiwavelength anomalous diffraction and refined it to 1.5 Å resolution. The present structure reveals an Hsp40-like peptide-binding domain, an Atx1-like metal-binding domain, and a flexible C terminus. Three metal-binding sites per dimer, defined by structural analysis of Cu-H144*UreE, are on the opposite face of the Atx1-like domain than observed in the copper metallochaperone. One metal bridges the two subunits via the pair of His-96 residues, whereas the other two sites involve metal coordination by His-110 and His-112 within each subunit. In contrast to the copper metallochaperone mechanism involving thiol ligand exchanges between structurally similar chaperones and target proteins, we propose that the Hsp40-like module interacts with urease apoprotein and/or other urease accessory proteins, while the Atx1-like domain delivers histidyl-bound nickel to the urease active site.Urease (EC 3.5.1.5) is a nickel-containing enzyme that catalyzes the hydrolysis of urea to produce ammonia and carbamate (1). Increased pH arising from this reaction is critical to the virulence of several human and animal pathogens (2). The crystal structure of urease from Klebsiella aerogenes provided the first three-dimensional model of the protein and revealed a unique dinuclear active site with the metal ions bridged by a carbamylated lysine residue (3, 4). Subsequent investigations of ureases from Bacillus pasteurii (5) and Helicobacter pylori (6) show essentially identical active site structures. Proper assembly of this metallocenter is a key step for maturation of urease and, in K. aerogenes, involves the products of the ureD,

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Section: Resultssupporting

confidence: 75%

Section: Resultssupporting

confidence: 59%

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

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Crystal Structure of Klebsiella aerogenesUreE, a Nickel-binding Metallochaperone for Urease Activation

Song

Mulrooney

Huber

et al. 2001

Journal of Biological Chemistry

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“…Superposition of the structures (19) have been shown to play a key role in metal binding by these proteins. In addition, the H96A mutation of an equivalent histidine in KaUreE significantly reduced urease activity in bacteria harboring this mutant (11,31). Similarly, the H102K mutation in HpUreE had a deleterious effect on Ni 2þ and Zn 2þ binding, based on ITC analysis of this mutant (4).…”

Section: Resultsmentioning

confidence: 99%

Crystal Structures of Apo and Metal-Bound Forms of the UreE Protein from Helicobacter pylori: Role of Multiple Metal Binding Sites,

et al. 2010

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Received March 10, 2010; Revised Manuscript Received July 15, 2010 ABSTRACT: The crystal structure of the urease maturation protein UreE from Helicobacter pylori has been determined in its apo form at 2.1 Å resolution, bound to Cu 2þ at 2.7 Å resolution, and bound to Ni 2þ at 3.1 Å resolution. Apo UreE forms dimers, while the metal-bound enzymes are arranged as tetramers that consist of a dimer of dimers associated around the metal ion through coordination by His102 residues from each subunit of the tetramer. Comparison of independent subunits from different crystal forms indicates changes in the relative arrangement of the N-and C-terminal domains in response to metal binding. The improved ability of engineered versions of UreE containing hexahistidine sequences at either the N-terminal or C-terminal end to provide Ni 2þ for the final metal sink (urease) is eliminated in the H102A version. Therefore, the ability of the improved Ni 2þ -binding versions to deliver more nickel is likely an effect of an increased local concentration of metal ions that can rapidly replenish transferred ions bound to His102.

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Urease

Hausinger¹,

Karplus²

2004

Handbook of Metalloproteins

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Urease catalyzes the hydrolysis of urea by all plants and many algae, fungi, and bacteria. The homohexameric enzyme from jack bean was crystallized in 1926 and shown to contain nickel in 1975, but the structure of the dinuclear metal site was first revealed in 1995 by using the heterotrimeric [(UreABC) 3 ] bacterial enzyme from Klebsiella aerogenes . A carbamylated lysine side chain and a water molecule bridge the nickel ions at each active site, with Ni‐1 also coordinated to two histidine side chains plus a terminal water and Ni‐2 bound to two histidines, one aspartate, and a terminal water. The kinetic properties and structures of various inhibited or mutant forms of the protein provide insights into the mechanism of urease catalysis.

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In Vivo and in Vitro Kinetics of Metal Transfer by the Klebsiella aerogenes Urease Nickel Metallochaperone, UreE

Abstract: The urease accessory protein encoded by ureE from Klebsiella aerogenes is proposed to deliver Ni(II) to the urease apoprotein during enzyme activation. Native UreE possesses a histidine-rich region at its carboxyl terminus that binds several equivalents of Ni

Cited by 60 publications

References 35 publications

Crystal Structure of Klebsiella aerogenesUreE, a Nickel-binding Metallochaperone for Urease Activation

Crystal Structure of Klebsiella aerogenesUreE, a Nickel-binding Metallochaperone for Urease Activation

Crystal Structures of Apo and Metal-Bound Forms of the UreE Protein from Helicobacter pylori: Role of Multiple Metal Binding Sites,

Urease

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