Probing ion binding sites in the Na+/Ca2+ exchanger.

Abstract:

:The membrane domain of the Na(+)/Ca(2+) exchanger (NCX) contains two conserved internal repeat sequences, designated the alpha-1 and alpha-2 repeats. We have studied the topological disposition of residues in the alpha repeats and a neighboring region by substituted cysteine accessibility scanning as well as the functional importance of these residues by kinetically evaluating transport activities of cysteine-substituted or other site-directed NCX1 mutants. The results suggest that the alpha-1 repeat contains a reentrant loop originating from extracellular side of the membrane, while the alpha-2 repeat and its neighboring region contain a complex reentrant loop structure originating from the cytoplasmic side. We identified several residues in the alpha-1 repeat loop whose mutation caused significant alterations in the interaction of NCX1 with a transport substrate Ca(2+)(o) and inhibitory ions Ni(2+) and Co(2+). On the other hand, we found residues in the alpha-2 repeat loop region whose mutation altered the interaction with an activating ion Li(+) and an inhibitory drug KB-R7943 in addition to the effect on Ca(2+)(o) and Ni(2+). Collectively, our data suggest that these alpha-repeat regions participate in the formation of ion translocation pathway of the exchanger and that the alpha-1 repeat loop plays an important role in ion selection.

journal_name

Ann N Y Acad Sci

authors

Shigekawa M,Iwamoto T,Uehara A,Kita S

doi

10.1111/j.1749-6632.2002.tb04710.x

subject

Has Abstract

pub_date

2002-11-01 00:00:00

pages

19-30

eissn

0077-8923

issn

1749-6632

journal_volume

976

pub_type

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