Differential Stabilization of the Three FMN Redox Forms by Tyrosine 94 and Tryptophan 57 in Flavodoxin from Anabaena and Its Influence on the Redox Potentials
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- 作者：Anabel Lostao ; Carlos Gó ; mez-Moreno ; Stephen G. Mayhew ; and Javier Sancho
- 刊名：Biochemistry
- 出版年：1997
- 出版时间：November 25, 1997
- 年：1997
- 卷：36
- 期：47
- 页码：14334 - 14344
- 全文大小：237K
- 年卷期：v.36,no.47(November 25, 1997)
- ISSN：1520-4995
文摘
Flavodoxins are electron transfer proteins that carry anoncovalently bound flavin mononucleotide molecule as the redox-active center. The redox potentials ofthe flavin nucleotide are profoundlyaltered upon interaction with the protein. In Anabaenaflavodoxin, as in many flavodoxins, the flavin issandwiched between two aromatic residues (Trp57 and Tyr94) thought tobe implicated in the alterationof the redox potentials. We have individually replaced these tworesidues by each of the other aromaticresidues, by alanine and by leucine. For each mutant, we havedetermined the redox potentials and thebinding energies of the oxidized FMN-apoflavodoxin complexes.From these data, the binding energiesof the semireduced and reduced complexes have been calculated.Comparison of the binding energies ofwild-type and mutant flavodoxins at the three redox states suggeststhat the interaction between Tyr94and FMN stabilizes the apoflavodoxin-FMN complex in all redox states.The oxidized and semireducedcomplexes are, however, more strongly stabilized than the reducedcomplex, making the semiquinone/hydroquinone midpoint potential more negative in flavodoxin than inunbound FMN. Trp57 also stabilizesall redox forms of FMN, thus cooperating with Tyr94 in strong FMNbinding. On the other hand, Trp57seems to slightly destabilize the semireduced complex relative to theoxidized one. Finally, we haveobserved that reduction of mutants lacking Trp57 is slow relative tothat of wild-type or mutants lackingTyr94, which suggests that Trp57 could play a role in the kinetics offlavodoxin redox reactions.