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

doi:10.1016/j.bbalip.2012.10.010

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

Hormel Institute

Research output: Contribution to journal › Article › peer-review

22 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 (λ_max ∼ 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 λ_max (∼ 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 language	English (US)
Pages (from-to)	417-427
Number of pages	11
Journal	Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids
Volume	1831
Issue number	2
DOIs	https://doi.org/10.1016/j.bbalip.2012.10.010
State	Published - Feb 2013

Bibliographical note

Funding Information:
We are grateful for the support by NIH/NIGMS GM45928 & GM34847 , NIH/NCI CA121493 , Spanish Ministerio de Ciencia e Innovacion (MICINN BFU2010-17711 ), Russian Foundation for Basic Research 012-04-00168 ; Abby Rockefeller Mauzé Trust , and the Dewitt Wallace, Maloris, Mayo, and Hormel Foundations . The molecular graphic images shown in Figs. 5, 7 , and S1 were produced using UCSF Chimera ( NIH P41 RR-01081 ) provided by the Resource for Biocomputing, Visualization, and Informatics at the University of California, San Francisco .

Keywords

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

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Kamlekar, R. K., Simanshu, D. K., Gao, Y.-G., Kenoth, R., Pike, H. M., Prendergast, F. G., Malinina, L., Molotkovsky, J. G., Venyaminov, S. Y., Patel, D. J., & Brown, R. E. (2013). The glycolipid transfer protein (GLTP) domain of phosphoinositol 4-phosphate adaptor protein-2 (FAPP2): Structure drives preference for simple neutral glycosphingolipids. Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids, 1831(2), 417-427. https://doi.org/10.1016/j.bbalip.2012.10.010

The glycolipid transfer protein (GLTP) domain of phosphoinositol 4-phosphate adaptor protein-2 (FAPP2): Structure drives preference for simple neutral glycosphingolipids. / Kamlekar, Ravi Kanth; Simanshu, Dhirendra K.; Gao, Yong-Guang et al.
In: Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids, Vol. 1831, No. 2, 02.2013, p. 417-427.

Research output: Contribution to journal › Article › peer-review

Kamlekar, RK, Simanshu, DK, Gao, Y-G, Kenoth, R, Pike, HM, Prendergast, FG, Malinina, L, Molotkovsky, JG, Venyaminov, SY, Patel, DJ & Brown, RE 2013, 'The glycolipid transfer protein (GLTP) domain of phosphoinositol 4-phosphate adaptor protein-2 (FAPP2): Structure drives preference for simple neutral glycosphingolipids', Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids, vol. 1831, no. 2, pp. 417-427. https://doi.org/10.1016/j.bbalip.2012.10.010

Kamlekar RK, Simanshu DK, Gao YG, Kenoth R, Pike HM, Prendergast FG et al. The glycolipid transfer protein (GLTP) domain of phosphoinositol 4-phosphate adaptor protein-2 (FAPP2): Structure drives preference for simple neutral glycosphingolipids. Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids. 2013 Feb;1831(2):417-427. doi: 10.1016/j.bbalip.2012.10.010

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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.",

keywords = "Divergent evolution, GLTP superfamily, Glycosphingolipid binding and transfer, Keywords, Membrane interaction, Near-UV and far-UV circular dichroism, Tryptophan fluorescence",

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

note = "Funding Information: We are grateful for the support by NIH/NIGMS GM45928 & GM34847 , NIH/NCI CA121493 , Spanish Ministerio de Ciencia e Innovacion (MICINN BFU2010-17711 ), Russian Foundation for Basic Research 012-04-00168 ; Abby Rockefeller Mauz{\'e} Trust , and the Dewitt Wallace, Maloris, Mayo, and Hormel Foundations . The molecular graphic images shown in Figs. 5, 7 , and S1 were produced using UCSF Chimera ( NIH P41 RR-01081 ) provided by the Resource for Biocomputing, Visualization, and Informatics at the University of California, San Francisco . ",

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AU - Simanshu, Dhirendra K.

AU - Gao, Yong-Guang

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

Abstract

Bibliographical note

Keywords

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