Biochemical and functional characterization of human phospholipid scramblase 4 (hPLSCR4)

Biol Chem. 2012 Oct;393(10):1173-81. doi: 10.1515/hsz-2012-0129.


Human phospholipid scramblase 4 (hPLSCR4), an isoform of the scramblase family, is a type II single-pass transmembrane protein whose function remains unknown. To understand its role, recombinant hPLSCR4 was obtained by cloning the ORF into a pET28 a(+) vector and overexpressed in Escherichia coli. Functional assay showed that Ca2+, Mg2+, and Zn2+ activate hPLSCR4 and mediate scrambling activity independent of the phospholipid head group. Far-UV-CD and fluorescence spectroscopy revealed that Ca2+ and Mg2+ binding induces conformation change in hPLSCR4, exposing hydrophobic patches of the protein, and Ca2+ has more affinity than Mg2+ and Zn2+. Stains-all studies further confirm that hPLSCR4 is a Ca2+-binding protein. Point mutation (Asp290→Ala) in hPLSCR4 decreased the Ca2+-binding affinity as well as Tb3+ luminescence, suggesting residues of the predicted Ca2+-binding motif are involved in Ca2+ binding. Functional reconstitution with (Asp290→Ala) mutant led to ~50% and ~40% decrease in scramblase activity in the presence of Ca2+ and Mg2+, respectively.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anilino Naphthalenesulfonates / chemistry
  • Calcium / metabolism
  • Calcium / pharmacology
  • Humans
  • Hydrophobic and Hydrophilic Interactions / drug effects
  • Mutagenesis, Site-Directed
  • Mutation
  • Phospholipid Transfer Proteins / chemistry*
  • Phospholipid Transfer Proteins / genetics
  • Phospholipid Transfer Proteins / isolation & purification
  • Phospholipid Transfer Proteins / metabolism*
  • Spectrometry, Fluorescence


  • 8-anilino-1-naphthalenesulfonic acid
  • Anilino Naphthalenesulfonates
  • PLSCR4 protein, human
  • Phospholipid Transfer Proteins
  • Calcium