The structure of urease activation complexes examined by flexibility analysis, mutagenesis, and small-angle X-ray scattering

Soledad Quiroz-Valenzuela; Sai Chetan K. Sukuru; Robert P. Hausinger; Leslie A. Kuhn; William T. Heller

doi:10.1016/j.abb.2008.09.004

The structure of urease activation complexes examined by flexibility analysis, mutagenesis, and small-angle X-ray scattering

Soledad Quiroz-Valenzuela
, Sai Chetan K. Sukuru
, Robert P. Hausinger
, Leslie A. Kuhn
, William T. Heller

Research output: Contribution to journal › Article › peer-review

35 Scopus citations

Abstract

Conformational changes of Klebsiella aerogenes urease apoprotein (UreABC)₃ induced upon binding of the UreD and UreF accessory proteins were examined by a combination of flexibility analysis, mutagenesis, and small-angle X-ray scattering (SAXS). ProFlex analysis of urease provided evidence that the major domain of UreB can move in a hinge-like motion to account for prior chemical cross-linking results. Rigidification of the UreB hinge region, accomplished through a G11P mutation, reduced the extent of urease activation, in part by decreasing the nickel content of the mutant enzyme, and by sequestering a portion of the urease apoprotein in a novel activation complex that includes all of the accessory proteins. SAXS analyses of urease, (UreABC-UreD)₃, and (UreABC-UreDF)₃ confirm that UreD and UreF bind near UreB at the periphery of the (UreAC)₃ structure. This study supports an activation model in which a domain-shifted UreB conformation in (UreABC-UreDF)₃ allows CO₂ and nickel ions to gain access to the nascent active site.

Original language	English
Pages (from-to)	51-57
Number of pages	7
Journal	Archives of Biochemistry and Biophysics
Volume	480
Issue number	1
DOIs	https://doi.org/10.1016/j.abb.2008.09.004
State	Published - Dec 1 2008

Funding

We thank Scott Mulrooney, Kimberly Anderson, and Yiming Mo for their assistance. This work was supported by the National Institutes of Health (DK45686 to R.P.H. and GM67249 to L.A.K.), the MSU Quantitative Biology and Modeling Initiative (Strategic Partnership Grant 71-4841), and Project KP1102010 of the Office of Biological and Environmental Research of the U.S. Department of Energy, under Contract No. DE-AC05-00OR22725 with Oak Ridge National Laboratory managed and operated by UT-Batelle, LLC.

Keywords

Activation
Flexibility
Small-angle X-ray scattering
Urease

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10.1016/j.abb.2008.09.004

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title = "The structure of urease activation complexes examined by flexibility analysis, mutagenesis, and small-angle X-ray scattering",

abstract = "Conformational changes of Klebsiella aerogenes urease apoprotein (UreABC)3 induced upon binding of the UreD and UreF accessory proteins were examined by a combination of flexibility analysis, mutagenesis, and small-angle X-ray scattering (SAXS). ProFlex analysis of urease provided evidence that the major domain of UreB can move in a hinge-like motion to account for prior chemical cross-linking results. Rigidification of the UreB hinge region, accomplished through a G11P mutation, reduced the extent of urease activation, in part by decreasing the nickel content of the mutant enzyme, and by sequestering a portion of the urease apoprotein in a novel activation complex that includes all of the accessory proteins. SAXS analyses of urease, (UreABC-UreD)3, and (UreABC-UreDF)3 confirm that UreD and UreF bind near UreB at the periphery of the (UreAC)3 structure. This study supports an activation model in which a domain-shifted UreB conformation in (UreABC-UreDF)3 allows CO2 and nickel ions to gain access to the nascent active site.",

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AU - Quiroz-Valenzuela, Soledad

AU - Sukuru, Sai Chetan K.

AU - Hausinger, Robert P.

AU - Kuhn, Leslie A.

AU - Heller, William T.

PY - 2008/12/1

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N2 - Conformational changes of Klebsiella aerogenes urease apoprotein (UreABC)3 induced upon binding of the UreD and UreF accessory proteins were examined by a combination of flexibility analysis, mutagenesis, and small-angle X-ray scattering (SAXS). ProFlex analysis of urease provided evidence that the major domain of UreB can move in a hinge-like motion to account for prior chemical cross-linking results. Rigidification of the UreB hinge region, accomplished through a G11P mutation, reduced the extent of urease activation, in part by decreasing the nickel content of the mutant enzyme, and by sequestering a portion of the urease apoprotein in a novel activation complex that includes all of the accessory proteins. SAXS analyses of urease, (UreABC-UreD)3, and (UreABC-UreDF)3 confirm that UreD and UreF bind near UreB at the periphery of the (UreAC)3 structure. This study supports an activation model in which a domain-shifted UreB conformation in (UreABC-UreDF)3 allows CO2 and nickel ions to gain access to the nascent active site.

AB - Conformational changes of Klebsiella aerogenes urease apoprotein (UreABC)3 induced upon binding of the UreD and UreF accessory proteins were examined by a combination of flexibility analysis, mutagenesis, and small-angle X-ray scattering (SAXS). ProFlex analysis of urease provided evidence that the major domain of UreB can move in a hinge-like motion to account for prior chemical cross-linking results. Rigidification of the UreB hinge region, accomplished through a G11P mutation, reduced the extent of urease activation, in part by decreasing the nickel content of the mutant enzyme, and by sequestering a portion of the urease apoprotein in a novel activation complex that includes all of the accessory proteins. SAXS analyses of urease, (UreABC-UreD)3, and (UreABC-UreDF)3 confirm that UreD and UreF bind near UreB at the periphery of the (UreAC)3 structure. This study supports an activation model in which a domain-shifted UreB conformation in (UreABC-UreDF)3 allows CO2 and nickel ions to gain access to the nascent active site.

KW - Activation

KW - Flexibility

KW - Small-angle X-ray scattering

KW - Urease

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ER -

The structure of urease activation complexes examined by flexibility analysis, mutagenesis, and small-angle X-ray scattering

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