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Analysis of a Soluble (UreD:UreF:UreG)2 Accessory Protein Complex and Its Interactions with Klebsiella aerogenes Urease by Mass Spectrometry |
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| Authors: | Mark A Farrugia Linjie Han Yueyang Zhong Jodi L Boer Brandon T Ruotolo Robert P Hausinger |
| Institution: | 1. Department of Biochemistry and Molecular Biology, Michigan State University, East Lansing, MI, 48824, USA 3. Department of Chemistry, University of Michigan, Ann Arbor, MI, 48109, USA 4. Department of Translational Science and Molecular Medicine, Michigan State University College of Human Medicine, Grand Rapids, MI, 49503, USA 2. Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, MI, 48824, USA |
| Abstract: | Maturation of the nickel-containing urease of Klebsiella aerogenes is facilitated by the UreD, UreF, and UreG accessory proteins along with the UreE metallo-chaperone. A fusion of the maltose binding protein and UreD (MBP-UreD) was co-isolated with UreF and UreG in a soluble complex possessing a (MBP-UreD:UreF:UreG)2 quaternary structure. Within this complex a UreF:UreF interaction was identified by chemical cross-linking of the amino termini of its two UreF protomers, as shown by mass spectrometry of tryptic peptides. A pre-activation complex was formed by the interaction of (MBP-UreD:UreF:UreG)2 and urease. Mass spectrometry of intact protein species revealed a pathway for synthesis of the urease pre-activation complex in which individual hetero-trimer units of the (MBP-UreD:UreF:UreG)2 complex bind to urease. Together, these data provide important new insights into the structures of protein complexes associated with urease activation. ? |
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