JBC

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Aliverti, A.
Right arrow Articles by Zanetti, G.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Aliverti, A.
Right arrow Articles by Zanetti, G.
Social Bookmarking
Add to CiteULike   Add to Complore   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati  
What's this?

J Biol Chem, Vol. 273, Issue 51, 34008-34015, December 18, 1998

Probing the Function of the Invariant Glutamyl Residue 312 in Spinach Ferredoxin-NADP+ Reductase

Alessandro AlivertiDagger , Zhan Deng§, Daniela RavasiDagger , Luciano PiubelliDagger , P. Andrew Karplus§, and Giuliana ZanettiDagger

From the Dagger  Dipartimento di Fisiologia e Biochimica Generali, Università degli Studi di Milano, Via Celoria 26, I-20133 Milano, Italy and the § Section of Biochemistry, Molecular and Cell Biology, Division of Biological Sciences, Cornell University, Ithaca, New York 14853

Ferredoxin-NADP+ reductase, the prototype of a large family of structurally related flavoenzymes, pairs single electrons carried by ferredoxin I and transfers them as a hydride to NADP+. Four mutants of the enzyme, in which Glu-312 was replaced with Asp, Gln, Leu, and Ala to probe the role of the residue charge, size, and polarity in the enzyme activity, have been heterologously expressed, purified, and characterized through steady-state, rapid kinetic studies, ligand-binding experiments, and three-dimensional structure determination by x-ray crystallography. The E312L mutant was the only one that was almost inactive (~1%), whereas unexpectedly the E312A reductase was 10-100% active with the various acceptors tested. Rapid kinetic absorption spectroscopy studies demonstrated that flavin reduction by NADPH was impaired in the mutants. Furthermore, NADP(H) binding was partially perturbed. These functional and structural studies lead us to conclude that Glu-312 does not fulfil the role of proton donor during catalysis, but it is required for proper binding of the nicotinamide ring of NADP(H). In addition, its charge modulates the two one-electron redox potentials of the flavin to stabilize the semiquinone form.

Copyright © 1998 by The American Society for Biochemistry and Molecular Biology, Inc.
Add to CiteULike CiteULike   Add to Complore Complore   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us   Add to Digg Digg   Add to Reddit Reddit   Add to Technorati Technorati    What's this?


This article has been cited by other articles:


Home page
 J. Biol. Chem.Home page
R. P. Ilagan, M. Tiso, D. W. Konas, C. Hemann, D. Durra, R. Hille, and D. J. Stuehr
Differences in a Conformational Equilibrium Distinguish Catalysis by the Endothelial and Neuronal Nitric-oxide Synthase Flavoproteins
J. Biol. Chem., July 11, 2008; 283(28): 19603 - 19615.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
K. Panda, S. Adak, D. Konas, M. Sharma, and D. J. Stuehr
A Conserved Aspartate (Asp-1393) Regulates NADPH Reduction of Neuronal Nitric-oxide Synthase: IMPLICATIONS FOR CATALYSIS
J. Biol. Chem., April 30, 2004; 279(18): 18323 - 18333.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
J. Tejero, M. Martinez-Julvez, T. Mayoral, A. Luquita, J. Sanz-Aparicio, J. A. Hermoso, J. K. Hurley, G. Tollin, C. Gomez-Moreno, and M. Medina
Involvement of the Pyrophosphate and the 2'-Phosphate Binding Regions of Ferredoxin-NADP+ Reductase in Coenzyme Specificity
J. Biol. Chem., December 5, 2003; 278(49): 49203 - 49214.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
L. Piubelli, A. Aliverti, A. K. Arakaki, N. Carrillo, E. A. Ceccarelli, P. A. Karplus, and G. Zanetti
Competition between C-terminal Tyrosine and Nicotinamide Modulates Pyridine Nucleotide Affinity and Specificity in Plant Ferredoxin-NADP+ Reductase
J. Biol. Chem., March 31, 2000; 275(14): 10472 - 10476.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
M. Medina, A. Luquita, J. Tejero, J. Hermoso, T. Mayoral, J. Sanz-Aparicio, K. Grever, and C. Gomez-Moreno
Probing the Determinants of Coenzyme Specificity in Ferredoxin-NADP+ Reductase by Site-directed Mutagenesis
J. Biol. Chem., April 6, 2001; 276(15): 11902 - 11912.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
M. Martinez-Julvez, I. Nogues, M. Faro, J. K. Hurley, T. B. Brodie, T. Mayoral, J. Sanz-Aparicio, J. A. Hermoso, M. T. Stankovich, M. Medina, et al.
Role of a Cluster of Hydrophobic Residues Near the FAD Cofactor in Anabaena PCC 7119 Ferredoxin-NADP+ Reductase for Optimal Complex Formation and Electron Transfer to Ferredoxin
J. Biol. Chem., July 13, 2001; 276(29): 27498 - 27510.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
A. K. Arakaki, E. G. Orellano, N. B. Calcaterra, J. Ottado, and E. A. Ceccarelli
Involvement of the Flavin si-Face Tyrosine on the Structure and Function of Ferredoxin-NADP+ Reductases
J. Biol. Chem., November 21, 2001; 276(48): 44419 - 44426.
[Abstract] [Full Text] [PDF]


HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
 All ASBMB Journals   Molecular and Cellular Proteomics 
 Journal of Lipid Research   ASBMB Today 
Copyright © 1998 by the American Society for Biochemistry and Molecular Biology.