The glycolipid transfer protein (GLTP) domain of phosphoinositol 4-phosphate adaptor protein-2 (FAPP2): Structure drives preference for simple neutral glycosphingolipids

Ravi Kanth Kamlekar, Dhirendra K. Simanshu, Yong Guang Gao, Roopa Kenoth, Helen M. Pike, Franklyn G. Prendergast, Lucy Malinina, Julian G. Molotkovsky, Sergei Yu Venyaminov, Dinshaw J. Patel, Rhoderick E. Brown

Research output: Contribution to journalArticlepeer-review

17 Scopus citations

Abstract

Phosphoinositol 4-phosphate adaptor protein-2 (FAPP2) plays a key role in glycosphingolipid (GSL) production using its C-terminal domain to transport newly synthesized glucosylceramide away from the cytosol-facing glucosylceramide synthase in the cis-Golgi for further anabolic processing. Structural homology modeling against human glycolipid transfer protein (GLTP) predicts a GLTP-fold for FAPP2 C-terminal domain, but no experimental support exists to warrant inclusion in the GLTP superfamily. Here, the biophysical properties and glycolipid transfer specificity of FAPP2-C-terminal domain have been characterized and compared with other established GLTP-folds. Experimental evidence for a GLTP-fold includes: i) far-UV circular dichroism (CD) showing secondary structure with high alpha-helix content and a low thermally-induced unfolding transition (∼ 41 C); ii) near-UV-CD indicating only subtle tertiary conformational change before/after interaction with membranes containing/lacking glycolipid; iii) Red-shifted tryptophan (Trp) emission wavelength maximum (λmax ∼ 352 nm) for apo-FAPP2-C-terminal domain consistent with surface exposed intrinsic Trp residues; iv) 'signature' GLTP-fold Trp fluorescence response, i.e., intensity decrease (∼ 30%) accompanied by strongly blue-shifted λmax (∼ 14 nm) upon interaction with membranes containing glycolipid, supporting direct involvement of Trp in glycolipid binding and enabling estimation of partitioning affinities. A structurally-based preference for other simple uncharged GSLs, in addition to glucosylceramide, makes human FAPP2-GLTP more similar to fungal HET-C2 than to plant AtGLTP1 (glucosylceramide-specific) or to broadly GSL-selective human GLTP. These findings along with the distinct mRNA exon/intron organizations originating from single-copy genes on separate human chromosomes suggest adaptive evolutionary divergence by these two GLTP-folds.

Original languageEnglish (US)
Pages (from-to)417-427
Number of pages11
JournalBiochimica et Biophysica Acta - Molecular and Cell Biology of Lipids
Volume1831
Issue number2
DOIs
StatePublished - Feb 2013

Keywords

  • Divergent evolution
  • GLTP superfamily
  • Glycosphingolipid binding and transfer
  • Keywords
  • Membrane interaction
  • Near-UV and far-UV circular dichroism
  • Tryptophan fluorescence

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology

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