Deconstructing the DGAT1 enzyme: Binding sites and substrate interactions


Autoria(s): Lopes, Jose L. S.; Nobre, Thatyane M.; Cilli, Eduardo M.; Beltramini, Leila M.; Araujo, Ana P. U.; Wallace, B. A.
Contribuinte(s)

Universidade Estadual Paulista (UNESP)

Data(s)

18/03/2015

18/03/2015

01/12/2014

Resumo

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Processo FAPESP: 09/17698-5

Diacylglycerol acyltransferase 1 (DGAT1) is a microsomal membrane enzyme responsible for the final step in the synthesis of triacylglycerides. Although DGATs from a wide range of organisms have nearly identical sequences, there is little structural information available for these enzymes. The substrate binding sites of DGAT1 are predicted to be in its large luminal extramembranous loop and to include common motifs with acyl-CoA cholesterol acyltransferase enzymes and the diacylglycerol binding domain found in protein kinases.In this study, synthetic peptides corresponding to the predicted binding sites of DGAT1 enzyme were examined using synchrotron radiation circular dichroism spectroscopy, fluorescence emission and adsorption onto lipid monolayers to determine their interactions with substrates associated with triacylglyceride synthesis (oleoyl-CoA and dioleoylglycerol). One of the peptides, Sit1, which includes the FYxDWWN motif common to both DGAT1 and acyl-CoA cholesterol acyltransferase, changes its conformation in the presence of both substrates, suggesting its capability to bind their acyl chains. The other peptide (Sit2), which includes the putative diacylglycerol binding domain HKWCIRHFYKP found in protein kinase C and diacylglycerol kinases, appears to interact with the charged headgroup region of the substrates. Moreover, in an extended-peptide which contains Sit1 and Sit2 sequences separated by a flexible linker, larger conformational changes were induced by both substrates, suggesting that the two binding sites may bring the substrates into close proximity within the membrane, thus catalyzing the formation of the triacylglyceride product. (C) 2014 Elsevier B.V. All Lights reserved.

Formato

3145-3152

Identificador

http://dx.doi.org/10.1016/j.bbamem.2014.08.017

Biochimica Et Biophysica Acta-biomembranes. Amsterdam: Elsevier Science Bv, v. 1838, n. 12, p. 3145-3152, 2014.

0005-2736

http://hdl.handle.net/11449/116446

10.1016/j.bbamem.2014.08.017

WOS:000343847200018

Idioma(s)

eng

Publicador

Elsevier B.V.

Relação

Biochimica Et Biophysica Acta-biomembranes

Direitos

closedAccess

Palavras-Chave #Diacylglycerol acyltransferase #Enzyme catalysis #Peptide-lipid interaction #Langmuir monolayer #Synchrotron radiation circular dichroism (SRCD) spectroscopy #Triglyceride synthesis
Tipo

info:eu-repo/semantics/article