Structural insights into triglyceride storage mediated by fat storage-inducing transmembrane (FIT) protein 2

David A. Gross, Erik L. Snapp, David L. Silver

Research output: Contribution to journalArticle

37 Citations (Scopus)

Abstract

Fat storage-Inducing Transmembrane proteins 1 & 2 (FIT1/FITM1 and FIT2/FITM2) belong to a unique family of evolutionarily conserved proteins localized to the endoplasmic reticulum that are involved in triglyceride lipid droplet formation. FIT proteins have been shown to mediate the partitioning of cellular triglyceride into lipid droplets, but not triglyceride biosynthesis. FIT proteins do not share primary sequence homology with known proteins and no structural information is available to inform on the mechanism by which FIT proteins function. Here, we present the experimentallysolved topological models for FIT1 and FIT2 using N-glycosylation site mapping and indirect immunofluorescence techniques. These methods indicate that both proteins have six-transmembrane-domains with both N- and C-termini localized to the cytosol. Utilizing this model for structure-function analysis, we identified and characterized a gain-offunction mutant of FIT2 (FLL(157-9)AAA) in transmembrane domain 4 that markedly augmented the total number and mean size of lipid droplets. Using limited-trypsin proteolysis we determined that the FLL(157-9)AAA mutant has enhanced trypsin cleavage at K86 relative to wild-type FIT2, indicating a conformational change. Taken together, these studies indicate that FIT2 is a 6 transmembrane domain-containing protein whose conformation likely regulates its activity in mediating lipid droplet formation.

Original languageEnglish (US)
Article numbere10796
JournalPLoS One
Volume5
Issue number5
DOIs
StatePublished - 2010

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transmembrane proteins
Triglycerides
Fats
triacylglycerols
lipids
droplets
Proteins
Lipids
Trypsin
trypsin
Protein Conformation
Proteolysis
Glycosylation
protein conformation
mutants
Indirect Fluorescent Antibody Technique
Sequence Homology
proteins
Biosynthesis
Endoplasmic Reticulum

ASJC Scopus subject areas

  • Agricultural and Biological Sciences(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Medicine(all)

Cite this

Structural insights into triglyceride storage mediated by fat storage-inducing transmembrane (FIT) protein 2. / Gross, David A.; Snapp, Erik L.; Silver, David L.

In: PLoS One, Vol. 5, No. 5, e10796, 2010.

Research output: Contribution to journalArticle

Gross, David A. ; Snapp, Erik L. ; Silver, David L. / Structural insights into triglyceride storage mediated by fat storage-inducing transmembrane (FIT) protein 2. In: PLoS One. 2010 ; Vol. 5, No. 5.
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