INVESTIGADORES
GONZALEZ Daniel Hector
artículos
Título:
Stereoselectivity of the interaction of E- and Z-2-phosphoenolbutyrate with maize leaf phosphoenolpyruvate carboxylase
Autor/es:
GONZALEZ, DANIEL H.; ANDREO, CARLOS S.
Revista:
EUROPEAN JOURNAL OF BIOCHEMISTRY
Editorial:
Elsevier
Referencias:
Año: 1988 vol. 173 p. 339 - 343
ISSN:
0014-2956
Resumen:
The aim of this work was to investigate the stereoselectivity of maize
leaf phosphoenolpyruvate carboxylase with E- and Z-2-phosphoenolbutyrate
as inhibitors and substrates. In addition, a procedure is presented for
the separation of the isomers of 2-phosphoenolbutyrate. The method is
based on the different interaction of those compounds with a strong
anion-exchange high-pressure liquid chromatography column using 50 mM
potassium phosphate (pH 3) as elution buffer, and allows the obtention
of pure E- and Z-P-enolbutyrate with high yield. The same system was
used to identify Z-P-enolbutyrate as the product of the phosphorylation
of 2-oxobutyrate by rabbit muscle pyruvate kinase. In the presence of 5
mM Mg2+, both isomers of P-enolbutyrate inhibited C4-plant
P-enolpyruvate carboxylase; the values of Ki were 15-20 microM and
100-110 microM for Z- and E-P-enolbutyrate, respectively. With 0.5 mM
Mn2+, the Z isomer was also effective as inhibitor (Ki = 35-40 microM),
while the E isomer produced activation of the carboxylase probably due
to its binding at an allosteric site. Both compounds were substrates of
the enzyme with similar V/Km values; however, V and Km for the two
isomers were significantly different (i.e. Km = 110 microM for
Z-P-enolbutyrate and 220 microM for E-P-enolbutyrate). The results
indicate the existence of stereoselectivity for the binding of
P-enolbutyrate to the active site of P-enolpyruvate carboxylase.
However, this fact does not affect the use of the isomers as substrates
by the plant carboxylase.