NICOLA JUAN PABLO
Congresos y reuniones científicas
Título:
Single amino acid substitutions change the sodium/iodide symporter (NIS) selectivity and stoichiometry
Autor/es:
PARODER-BELENITSKY, M; MAESTRAS, M; DOHAN, O; NICOLA, JP; REYNA-NEYRA, A; FOLLENZI, A; ESKANDARI, S; AMZEL, LM; CARRASCO, N
Lugar:
San Fransisco
Reunión:
Congreso; Biophysical Society 54th Annual Meeting; 2010
Institución organizadora:
Biophysical Society
Resumen:
The Na+/I- symporter (NIS) is a key plasma membrane protein that mediates
active I- transport in the thyroid and such other tissues as salivary glands, stomach,
and lactating breast. NIS-mediated I- uptake is the first step in thyroid hormone
biosynthesis. NIS mediates the inward simultaneous movement of Na+
and I- with a 2:1 stoichiometry, thus resulting in a net transfer of positive charge
into the cell (i.e., electrogenic transport). We recently reported that NIS translocates
different anion substrates with different stoichiometries, as Na+/perchlorate
(or perrhenate) transport is electroneutral. Valuable mechanistic information
on NIS has been obtained by the characterization of NIS mutants that
cause congenital I- transport defect in patients. Here we provide a detailed study
of the G93R NIS mutant. As we substituted neutral amino acids at this position,
we observed that the longer the side chain of the substituted residue, the lower
the protein’s activity. G93T and G93N NIS exhibited significantly higher Km
values for I- than WT NIS, the first time that such a change has been observed
in any NIS mutants. Strikingly, we show by kinetic analysis that G93T-mediated
Na+/perrhenate symport is electrogenic with a 2:1 stoichiometry, a discovery
confirmed by the detection of currents elicited by perrhenate (or perchlorate)
in G93T NIS-expressing X. laevis oocytes in electrophysiological
experiments. These observations demonstrate that a single amino acid substitution
at position 93 converts NIS-mediated Na+/perchlorate (or perrhenate)
transport stoichiometry from electroneutral to electrogenic. Based on the 3-D
structure of the bacterial Na+/galactose transporter, we built a 3-D homology
model of NIS and we propose a mechanism in which changes from an outwardly
open to an inwardly open conformation during the transport cycle use
G93 as a pivot.
active I- transport in the thyroid and such other tissues as salivary glands, stomach,
and lactating breast. NIS-mediated I- uptake is the first step in thyroid hormone
biosynthesis. NIS mediates the inward simultaneous movement of Na+
and I- with a 2:1 stoichiometry, thus resulting in a net transfer of positive charge
into the cell (i.e., electrogenic transport). We recently reported that NIS translocates
different anion substrates with different stoichiometries, as Na+/perchlorate
(or perrhenate) transport is electroneutral. Valuable mechanistic information
on NIS has been obtained by the characterization of NIS mutants that
cause congenital I- transport defect in patients. Here we provide a detailed study
of the G93R NIS mutant. As we substituted neutral amino acids at this position,
we observed that the longer the side chain of the substituted residue, the lower
the protein’s activity. G93T and G93N NIS exhibited significantly higher Km
values for I- than WT NIS, the first time that such a change has been observed
in any NIS mutants. Strikingly, we show by kinetic analysis that G93T-mediated
Na+/perrhenate symport is electrogenic with a 2:1 stoichiometry, a discovery
confirmed by the detection of currents elicited by perrhenate (or perchlorate)
in G93T NIS-expressing X. laevis oocytes in electrophysiological
experiments. These observations demonstrate that a single amino acid substitution
at position 93 converts NIS-mediated Na+/perchlorate (or perrhenate)
transport stoichiometry from electroneutral to electrogenic. Based on the 3-D
structure of the bacterial Na+/galactose transporter, we built a 3-D homology
model of NIS and we propose a mechanism in which changes from an outwardly
open to an inwardly open conformation during the transport cycle use
G93 as a pivot.
