Chaperone-assisted cryo-EM structure of P. aeruginosa PhuR reveals molecular basis for heme binding.
| Autor: | Knejski PP; Department of Biochemistry and Molecular Biology, The University of Chicago, Chicago, IL 60637, USA; Laboratory of Medical Biology, Faculty of Biotechnology, University of Wrocław, 50-383 Wrocław, Poland., Erramilli SK; Department of Biochemistry and Molecular Biology, The University of Chicago, Chicago, IL 60637, USA. Electronic address: satchal@gmail.com., Kossiakoff AA; Department of Biochemistry and Molecular Biology, The University of Chicago, Chicago, IL 60637, USA; Institute for Biophysical Dynamics, The University of Chicago, Chicago, IL 60637, USA. Electronic address: koss@bsd.uchicago.edu. |
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| Jazyk: | angličtina |
| Zdroj: | Structure (London, England : 1993) [Structure] 2024 Apr 04; Vol. 32 (4), pp. 411-423.e6. Date of Electronic Publication: 2024 Feb 06. |
| DOI: | 10.1016/j.str.2024.01.007 |
| Abstrakt: | Pathogenic bacteria, such as Pseudomonas aeruginosa, depend on scavenging heme for the acquisition of iron, an essential nutrient. The TonB-dependent transporter (TBDT) PhuR is the major heme uptake protein in P. aeruginosa clinical isolates. However, a comprehensive understanding of heme recognition and TBDT transport mechanisms, especially PhuR, remains limited. In this study, we employed single-particle cryogenic electron microscopy (cryo-EM) and a phage display-generated synthetic antibody (sAB) as a fiducial marker to enable the determination of a high-resolution (2.5 Å) structure of PhuR with a bound heme. Notably, the structure reveals iron coordination by Y529 on a conserved extracellular loop, shedding light on the role of tyrosine in heme binding. Biochemical assays and negative-stain EM demonstrated that the sAB specifically targets the heme-bound state of PhuR. These findings provide insights into PhuR's heme binding and offer a template for developing conformation-specific sABs against outer membrane proteins (OMPs) for structure-function investigations. Competing Interests: Declaration of interests The authors declare no competing interests. (Copyright © 2024 Elsevier Ltd. All rights reserved.) |
| Databáze: | MEDLINE |
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