Benzophenone-containing cholesterol surrogates

Spencer, Thomas A.; Wang, Pingzhen; Russel, Jonathon S.; Blank, David H.A.; Huuskonen, Jarkko; Fielding, Phoebe E.; Fielding, Christopher J.

doi:10.1194/jlr.m400081-jlr200

Cited by 22 publications

(31 citation statements)

References 61 publications

(63 reference statements)

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“…Effect of SUV lipid composition on the α-helical content of proSCP-2 and SCP-2 N-terminal peptides upon membrane interaction. Circular dichroism spectra were obtained as described in Experimental Procedures: (A and B) peptide [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20] Pro, (C and D) peptide Pro, (E and F) peptide Colocalization of Cy5-SCP-2 and Cy5-proSCP-2 with a plasma membrane marker in L cells. L cells were incubated in the presence of Cy5-SCP-2 (A-D) or Cy5-proSCP-2 (E-H) and the plasma membrane marker, AlexaFluor 488-cholera toxin B (AF488-CTB), and examined by laser scanning confocal microscopy (LSCM) as described in Experimental Procedures.…”

Section: Discussionmentioning

confidence: 99%

“…A 41-kDa fragment of peroxin 5 [Pex5p(C)] comprising residues 235-602 and encompassing the sevenpredicted tetratricopeptide repeat (TPR) region as well as the peroxisomal targeting signal-1 (PTS-1) binding region of human Pex5p was generated and purified by J. M. Berg (National Institutes of Health) (18). Free cholesterol benzophenone (FCBP) was synthesized by T. A. Spencer (Department of Chemistry, Dartmouth College) (19). Radiolabelled free cholesterol benzophenone ([ 3 H]FCBP) was prepared for our laboratory by Moravek Biochemicals (Brea, CA) utilizing a procedure (20) that incorporates 3 H at position C-3 of the cholesterol moiety.…”

Section: Experimental Procedures Materialsmentioning

confidence: 99%

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Structure and Function of the Sterol Carrier Protein-2 N-Terminal Presequence

et al. 2008

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Although sterol carrier protein-2 (SCP-2) is encoded as a precursor protein (proSCP-2), little is known regarding the structure and function of the 20-amino acid N-terminal presequence. As shown herein, the presequence contains significant secondary structure and alters SCP-2: (i) secondary structure (CD), (ii) tertiary structure (aqueous exposure of Trp shown by UV absorbance, fluorescence, fluorescence quenching), (iii) ligand binding site [Trp response to ligands, peptide cross-linked by photoactivatable free cholesterol (FCBP)], (iv) selectivity for interaction with anionic phospholipid-rich membranes, (v) interaction with a peroxisomal import protein [FRET studies of Pex5p(C) binding], the N-terminal presequence increased SCP-2's affinity for Pex5p(C) by 10-fold, and (vi) intracellular targeting in living and fixed cells (confocal microscopy). Nearly 5-fold more SCP-2 than proSCP-2 colocalized with plasma membrane lipid rafts/caveolae (AF488-CTB), 2.8-fold more SCP-2 than proSCP-2 colocalized with a mitochondrial marker (Mitotracker), but nearly 2-fold less SCP-2 than proSCP-2 colocalized with peroxisomes (AF488-antibody to PMP70). These data indicate the importance of the N-terminal presequence in regulating SCP-2 structure, cholesterol localization within the ligand binding site, membrane association, and, potentially, intracellular targeting.Keywords sterol carrier protein-2; presequence; cholesterol; peroxin; peroxisome Sterol carrier protein-2 (SCP-2) 1 is a ubiquitous, soluble protein present at highest level in tissues involved in cholesterol uptake (intestine), oxidation (liver, steroidogenic cells), and/or elimination (liver) (reviewed in ref 1). SCP-2 exhibits high (nanomolar K d values) affinity for

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Section: Discussionmentioning

confidence: 99%

Section: Experimental Procedures Materialsmentioning

confidence: 99%

Structure and Function of the Sterol Carrier Protein-2 N-Terminal Presequence

et al. 2008

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“…We have been preparing benzophenone-containing analogues of sterols 1 and phospholipids 2 for use as photoaffinity labelling agents in studies of cellular cholesterol efflux and HDL formation. [3][4][5] In particular, the cholesterol analogue 1, known as FCBP, 3 and the fatty acid analogue 2 ( Figure 1), 2 were required in radioactive form for use as biochemical research tools.…”

Section: Introductionmentioning

confidence: 99%

Preparation of isotopically labelled benzophenone-containing lipid analogues

Wang

Spencer

2005

J Label Compd Radiopharm

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“…Recently, the syntheses of benzophenone-containing PC analogues and of benzophenone-containing cholesterol surrogates have been reported. 43,44 The benzophenone-derivatized sterols were shown to compete with cholesterol for apolipoprotein A-I-induced cellular efflux. 44 3-Trifluoromethyl-3-phenyl diazirines (TPD) generate carbenes upon UV light exposure that have a life time in the order of nanoseconds.…”

Section: Photoactivatable Lipid Crosslinkers: Properties and Applicatmentioning

confidence: 99%

“…43,44 The benzophenone-derivatized sterols were shown to compete with cholesterol for apolipoprotein A-I-induced cellular efflux. 44 3-Trifluoromethyl-3-phenyl diazirines (TPD) generate carbenes upon UV light exposure that have a life time in the order of nanoseconds. 45 No intramolecular rearrangements are observed and they insert in any type of bond, including non-reactive C-H bonds.…”

Section: Photoactivatable Lipid Crosslinkers: Properties and Applicatmentioning

confidence: 99%

Proteome-wide detection of phospholipid–protein interactions in mitochondria by photocrosslinking and click chemistry

Gubbens

Kroon

2010

Mol. BioSyst.

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Photoactivatable lipid analogues are uniquely suited for the detection of lipid-protein interactions in biological membranes. Based on photocrosslinking, new methodology has been developed for the proteome-wide detection of lipid-protein interactions. Bifunctional lipid analogues containing a tag for click chemistry in addition to the photoactivatable moiety enable the enrichment of the crosslinked proteins that is required for subsequent identification by mass spectrometry. In principle the phospholipid interaction-based membrane protein proteomics approach is applicable to any biomembrane and any lipid. Here, we review the background and the development of the new methodology. Results obtained with photocrosslinking in purified mitochondrial membranes from the yeast Saccharomyces cerevisiae are summarized and future perspectives discussed.

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Benzophenone-containing cholesterol surrogates

Cited by 22 publications

References 61 publications

Structure and Function of the Sterol Carrier Protein-2 N-Terminal Presequence

Structure and Function of the Sterol Carrier Protein-2 N-Terminal Presequence

Preparation of isotopically labelled benzophenone-containing lipid analogues

Proteome-wide detection of phospholipid–protein interactions in mitochondria by photocrosslinking and click chemistry

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