INVESTIGADORES
ADAMO hugo Pedro
congresos y reuniones científicas
Título:
The effects of mutations at Glu99 in the autoinhibition of the human plasma membrane Ca2+-pump (PMCA).
Autor/es:
MAZZITELLI LUCIANA ROMINA; ADAMO HUGO PEDRO
Lugar:
Salta, Argentina
Reunión:
Congreso; XXXIX Annual Meeting of Argentinean Biophysical Society SAB 2010, workshop CeBEM, structural biology in Latin America, 3rd Latin American Protein Society Meeting; 2010
Institución organizadora:
XXXIX Annual Meeting of Argentinean Biophysical Society SAB 2010, workshop CeBEM, structural biology in Latin America, 3rd Latin American Protein Society Meeting
Resumen:
The
PMCA belongs to the P2B family of P-ATPases, which are characterized their autoinhibitory
mechanism. The precise molecular mechanism responsible for the autoinhibition
of the pump is not known. It has been proposed that the PMCAs autoinhibition
involves the blockage of the active site of the enzyme by the rearrangement of
the N- and C-terminal segments of the molecule (1). The conformational change
leading to activation is triggered by the binding of Ca2+-calmodulin
to its site in the C-terminal segment. Glu99 is located in the
connecting stretch between the PMCAs A domain and transmembrane segment M1. Mutations
at the equivalent residue have been shown to activate the autoinhibited plant
Ca2+ pump ACA2 (2). We have
investigated the effect substituting Glu99 either by Gln or Lys. We
found that both mutants retained about 70% of the maximal activity of the wild
type enzyme. In the absence of calmodulin the activity of the wild type enzyme increased
with the concentration of Ca2+ with K1/2 of 1.3 uM
Ca2+ , and at saturating Ca2+ attained 65% of its maximal activity in the presence of
calmodulin. In the same experimental
conditions mutant Glu99Lys exhibited a slightly increased apparent
affinity for Ca2+ affinity K1/2 (1 uM Ca2+)
but at saturating Ca2+ attained
93% of its maximal activity in the
presence of calmodulin. In contrast the behavior of mutant Glu99Gln was
near wild type. These results show that
substitution of Glu99 for a positively charged residue partially disrupts the
autoihbitition mainly by increasing the maximal activity of the autoinhibited
enzyme