Structural and functional analyses of PpENA1 provide insights into cation binding by type IID P-type ATPases in lower plants and fungi

Damian P. Drew, Maria Hrmova, Christina Lunde, Andrew K. Jacobs, Mark Tester, Geoffrey B. Fincher

Research output: Contribution to journalArticlepeer-review

6 Scopus citations

Abstract

PpENA1 is a membrane-spanning transporter from the moss Physcomitrella patens, and is the first type IID P-type ATPase to be reported in the plant kingdom. In Physcomitrella, PpENA1 is essential for normal growth under moderate salt stress, while in yeast, type IID ATPases provide a vital efflux mechanism for cells under high salt conditions by selectively transporting Na+ or K+ across the plasma membrane. To investigate the structural basis for cation-binding within the type IID ATPase subfamily, we used homology modeling to identify a highly conserved cation-binding pocket between membrane helix (MH) 4 and MH 6 of the membrane-spanning pore of PpENA1. Mutation of specific charged and polar residues on MHs 4-6 resulted in a decrease or loss of protein activity as measured by complementation assays in yeast. The E298S mutation on MH 4 of PpENA1 had the most significant effect on activity despite the presence of a serine at this position in fungal type IID ATPases. Activity was partially restored in an inactivated PpENA1 mutant by the insertion of two additional serine residues on MH 4 and one on MH 6 based on the presence of these residues in fungal type IID ATPases. Our results suggest that the residues responsible for cation-binding in PpENA1 are distinct from those in fungal type IID ATPases, and that a fungal-type cation binding site can be successfully engineered into the moss protein.

Original languageEnglish (US)
Pages (from-to)1483-1492
Number of pages10
JournalBiochimica et Biophysica Acta - Biomembranes
Volume1808
Issue number6
DOIs
StatePublished - Jun 1 2011

Keywords

  • Homology modeling
  • Na pump
  • Physcomitrella
  • PpENA1
  • Type IID ATPase

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Cell Biology

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