IBR 13079
INSTITUTO DE BIOLOGIA MOLECULAR Y CELULAR DE ROSARIO
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
The Ferredoxin NADP+ Reductase from E. coli contains NADP+ tightly bound
Autor/es:
MUSUMECI, M. A.; CATALANO DUPUY, D. L.; CECCARELLI, E. A.
Lugar:
Berkeley
Reunión:
Simposio; 17th International Symposium on Flavins and Flavoproteins; 2011
Institución organizadora:
Universidad de Berkeley
Resumen:
Ferredoxin-NADP+
reductases (FNRs, EC 1.18.1.2) are ubiquitous enzymes which function as switch
in redox reactions between NADPH and ferredoxin or flavodoxin. The capability
of FNRs to catalyse reactions between mono and
bi-electronic substrates is due to their prosthetic group FAD, which can exist
in different redox states. In plants, FNRs have evolved to produce high amounts
of NADPH during photosynthesis. However, the inverse reaction is favoured in
bacteria, namely the reduction of ferredoxin or flavodoxin from NADPH. In
addition, FNR in bacteria regulates the homeostasis of
the NADP(H) pool as a defensive response.
In this work we have performed a comparative study of
the NADP+ binding mode in pea FNR (a plastidic type FNR) and Escherichia
coli FNR (a bacterial type FNR). The structure of E. coli FNR mutant
in which the purification and crystallization protocols yielded an enzyme in
complex with NADP+ was previously reported (1). Through HPLC
analysis, we
have observed that a fraction of wild-type E. coli FNR molecules are
able to retain NADP+ tightly bound when the enzyme is expressed in E.
coli cells and during the purification procedure, at difference of pea FNR.
The bacterial FNR with NADP+ bound could be separated from the FNR
alone and differences in their absorbance spectra were observed. Studies
of NADP+ binding mode suggested that a conformational change may
slowly occur as a function of time.
In order to obtain more
information about the interaction of these FNRs with the nucleotide substrate,
the binding of P-AMP was analyzed. By comparing the obtained results for the
NADP+ and P-AMP interaction we found that the nicotinamide portion
contributes to the NADP+ binding in E. coli FNR, at
difference it does not occur in the case of the pea FNR.
The implications of this
particular NADP+ binding mode in the physiological function of E.
coli FNR are discussed.
