CEFOBI   05405
CENTRO DE ESTUDIOS FOTOSINTETICOS Y BIOQUIMICOS
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
Study of C4-organic acids binding to NAD-malic enzyme 1 from Arabidopsis mitochondria by quenching of protein native fluorescence
Autor/es:
MARCOS A. TRONCONI, MARIEL C. GERRARD WHEELER, MARÍA F. DRINCOVICH AND CARLOS S. ANDREO
Lugar:
Salta
Reunión:
Congreso; Latin American Protein Society Meeting; 2010
Institución organizadora:
Sociadad Argentina de Biofísica
Resumen:
Plant
mitochondria can use L-malate or fumarate, which accumulate at high levels in
plant cells, as respiratory substrates. In part, this plant property is due to
the presence of a mitochondrial NAD-dependent malic enzyme (NAD-ME) that
displays particular structural and regulatory characteristics. In Arabidopsis,
the enzyme acts as two homodimers (NAD-ME1 and NAD-ME2) and also can form a heterodimer1.
NAD-ME1 and -2 homodimers are differently regulated by metabolites of central
carbon pathways2. In this regard, while the NAD-ME1 activity is positively
modulated by fumarate or oxaloacetate, these C4-organic acids not substantially
modify the NAD-ME2 activity. In this work, the interaction of NAD-ME1 with fumarate
and oxaloacetate was investigated by quenching of tryptophan fluorescence
studies. The results indicated that while in NAD-ME2 the quenching process
arise from a collisional effect of these metabolites, in NAD-ME1 the
fluorescence decay can be explained by a static process that involve the
binding of quenchers to the protein surface. The characterization of NAD-ME1
mutant versions revealed that these acids would bind to a single allosteric site
on the enzyme. Moreover, the dissociation constants (Kd) values calculated
from NAD-ME1 quenching data, indicate that fumarate is the ligand that binds
with higher affinity. Finally, the temperature dependence of the thermodynamic
parameters (ΔG, ΔH and ΔS) suggests that the fumarate binding process is
entropicaly conducted and important alterations in the disposition of lateral
chains of aminoacids are produced by the ligand-protein association.