Dynamic Hydrogen-Bonding Network in the Distal Pocket of the Nitrosyl Complex of Pseudomonas aeruginosa cd Nitrite Reductase.
cd nitrite reductase (NIR) is a key enzyme in the denitrification process that reduces nitrite to nitric oxide (NO). It contains a specialized d-heme cofactor, found only in this class of enzymes, where the substrate, nitrite, binds and is converted to NO. For a long time, it was believed that NO mu...
| Published in: | Journal of the American Chemical Society Vol. 133; no. 9; pp. 3043 - 3056 |
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| Main Authors: | , , , , , |
| Format: | Article |
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American Chemical Society
3/9/2011
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| Online Access: | View this record in EBSCOhost |
| fields | @attributes: recordID: 1 pdfLink: plink: https://search.ebscohost.com/login.aspx?direct=true&db=hlh&AN=62251590&site=ehost-live header: @attributes: shortDbName: hlh uiTerm: 62251590 longDbName: Humanities International Complete uiTag: AN controlInfo: bkinfo: jinfo: jid: 00027863 ACS jtl: Journal of the American Chemical Society issn: 00027863 maglogo: N pubinfo: dt: 3/9/2011 vid: 133 iid: 9 pid: 997 pub: American Chemical Society artinfo: ui: 62251590 10.1021/ja109688w ppf: 3043 ppct: 13 formats: tig: atl: Dynamic Hydrogen-Bonding Network in the Distal Pocket of the Nitrosyl Complex of Pseudomonas aeruginosa cd Nitrite Reductase. aug: au: Radoul, Marina Bykov, Dmytro Rinaldo, Serena Cutruzzolà, Francesca Neese, Frank Goldfarb, Daniella affil: Department of Chemical Physics, Weizmann Institute of Science, Rehovot 76100, Israel Dipartimento di Scienze Biochimiche "A. Rossi Fanelli", Universit di Roma "La Sapienza", Rome, Italy Institute for Physical and Theoretical Chemistry, University of Bonn, Wegelerstrasse 12, Bonn -53115, Germany su: Hydrogen bonding Molecular association Pseudomonas aeruginosa Density functionals Nitrogen compounds Scission (Chemistry) sug: subj: Hydrogen bonding Molecular association Pseudomonas aeruginosa Density functionals Nitrogen compounds Scission (Chemistry) ab: cd nitrite reductase (NIR) is a key enzyme in the denitrification process that reduces nitrite to nitric oxide (NO). It contains a specialized d-heme cofactor, found only in this class of enzymes, where the substrate, nitrite, binds and is converted to NO. For a long time, it was believed that NO must be released from the ferric d-heme to avoid enzyme inhibition by the formation of ferrous-nitroso complex, which was considered as a dead-end product. However, recently an enhanced rate of NO dissociation from the ferrous form, not observed in standard b-type hemes, has been reported and attributed to the unique d-heme structure (Rinaldo, S.; Arcovito, A.; Brunori, M.; Cutruzzol, F. J. Biol. Chem. 2007, 282, 14761-14767). Here, we report on a detailed study of the spatial and electronic structure of the ferrous d-heme NO complex from Pseudomonas aeruginosa cd NIR and two mutants Y10F and H369A/H327A in solution, searching for the unique properties that are responsible for the relatively fast release. There are three residues at the "distal" side of the heme (Tyr, His, and His), and in this work we focus on the identification and characterization of possible H-bonds they can form with the NO, thereby affecting the stability of the complex. For this purpose, we have used high field pulse electron-nuclear double resonance (ENDOR) combined with density functional theory (DFT) calculations. The DFT calculations were essential for assigning and interpreting the ENDOR spectra in terms of geometric structure. We have shown that the NO in the nitrosyl d-heme complex of cd NIR forms H-bonds with Tyr and His, whereas the second conserved histidine, His, appears to be less involved in NO H-bonding. This is in contrast to the crystal structure that shows that Tyr is removed from the NO. We have also observed a larger solvent accessibility to the distal pocket in the mutants as compared to the wild-type. Moreover, it was shown that the H-bonding network within the active site is dynamic and that a change in the protonation state of one of the residues does affect the strength and position of the H-bonds formed by the others. In the Y10F mutant, His is closer to the NO, whereas mutation of both distal histidines displaces Tyr, removing its H-bond. The implications of the H-bonding network found in terms of the complex stability and catalysis are discussed. pubtype: Academic Journal doctype: Article src: R language: English refInfo: copyright: @attributes: flag: Y dt: @attributes: year: 2011 holdings: @attributes: islocal: N |
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