Document Detail


Cyanide and nitric oxide binding to reduced protocatechuate 3,4-dioxygenase: insight into the basis for order-dependent ligand binding by intradiol catecholic dioxygenases.
MedLine Citation:
PMID:  9369476     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
EPR-silent, chemically reduced protocatechuate 3,4-dioxygenase (Er) binds NO at the active site Fe2+ to yield an EPR-active, S = 3/2 species that blocks subsequent binding of all other exogenous ligands. In contrast, addition of NO to a preformed Er.CN- complex yields an EPR-active, S = 1/2 species [Er.(CN)x.NO] that exhibits resolved superhyperfine splitting from 13CN-, 15/14NO, and a protein-derived 14N. Simulations of the EPR spectra observed for the Er.(CN)x.NO complex formed with 12CN- and 13CN- (1:1) show that CN- binds in two iron ligand sites (x >/= 2). The two cyanides exhibit similar, but distinguishable, hyperfine coupling constants. This demonstrates unambiguously that at least three exogenous ligands (two cyanides and NO) can bind to the Fe2+ simultaneously and strongly suggests that at least one histidine ligand is retained in the complex. The Er.(CN)>/=2.NO complex readily exchanges both of the bound cyanides for the substrate analog, 2-hydroxyisonicotinic acid N-oxide (INO), to form a Er.INO.NO complex exhibiting the same S = 3/2 type EPR spectrum that is observed for this complex in the absence of CN-. Because the dead-end Er.NO complex does not accumulate during the exchange, the results suggest that Er.(CN)>/=2. NO and Er.INO.NO are in conformational states that allow facile exchange of INO and CN- but not NO. The results are interpreted in the context of the known X-ray crystal structures for the ferric form of the resting enzyme (Eox) and numerous Eox.substrate, inhibitor, and CN- complexes, all of which have a charge neutral iron center. It is proposed that the binding of one CN- causes dissociation of an anionic endogenous ligand which begins a series of conformational changes analogous to those initiated by anionic substrate binding to Eox. This results in a new unique coordination site for NO, and a new second site for CN-; both cyanide sites are utilized when the enzyme subsequently binds substrates or INO.
Authors:
A M Orville; J D Lipscomb
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  Biochemistry     Volume:  36     ISSN:  0006-2960     ISO Abbreviation:  Biochemistry     Publication Date:  1997 Nov 
Date Detail:
Created Date:  1997-12-19     Completed Date:  1997-12-19     Revised Date:  2007-11-14    
Medline Journal Info:
Nlm Unique ID:  0370623     Medline TA:  Biochemistry     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  14044-55     Citation Subset:  IM    
Affiliation:
Department of Biochemistry, Medical School, and Center for Metals in Biocatalysis, University of Minnesota, Minneapolis, Minnesota 55455-0347, USA.
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MeSH Terms
Descriptor/Qualifier:
Cyanides / chemistry,  metabolism*
Electron Spin Resonance Spectroscopy
Iron
Ligands
Models, Chemical
Nitric Oxide / chemistry,  metabolism*
Oxidation-Reduction
Protocatechuate-3,4-Dioxygenase / chemistry,  metabolism*
Grant Support
ID/Acronym/Agency:
GM 24689/GM/NIGMS NIH HHS; GM-07323/GM/NIGMS NIH HHS
Chemical
Reg. No./Substance:
0/Cyanides; 0/Ligands; 10102-43-9/Nitric Oxide; 7439-89-6/Iron; EC 1.13.11.3/Protocatechuate-3,4-Dioxygenase

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