Biochemical characterization of the lipid-binding properties of a broccoli cuticular wax-associated protein, WAX9D, and its application.
In this study, we showed that WAX9D, a nonspecific lipid-transfer protein found in broccoli, binds palmitate (C16) and stearate (C18) with dissociation constants of 0.56 muM and 0.52 muM, respectively. WAX9D was fused to thioredoxin protein by genetic manipulation to enhance its solubility. The data revealed strong interaction of Trx-WAX9D with palmitate and stearate. The dissociation constants of Trx-WAX9D for palmitate and stearate were 1.1 muM and 6.4 muM, respectively. The calculated number of binding sites for palmitate and stearate was 2.5 to 2.7, indicating that Trx-WAX9D can bind three molecules of fatty acids. Additionally, Trx-WAX9D was shown to inhibit the apoptotic effect of palmitate in endothelial cells. Our data using Trx-WAX9D provide insight into the broad spectrum of its biological applications with specific palmitate binding.
Ahn
Sunyoung
S
Department of Molecular Biology and Institute of Nanosensor and Biotechnology, BK21 Graduate Program for RNA Biology, Dankook University, Yongin 448-701, Korea.
Kim
Jongmin
J
Pyee
Jaeho
J
Park
Heonyong
H
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001937:Brassica / Q000737:chemistry* / Q000378:metabolism; D002352:Carrier Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism / Q000494:pharmacology; D002417:Cattle; D004353:Drug Evaluation, Preclinical; D042783:Endothelial Cells / Q000187:drug effects / Q000502:physiology; D005227:Fatty Acids / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins; D019308:Palmitic Acid / Q000494:pharmacology; D010940:Plant Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism / Q000494:pharmacology; D011485:Protein Binding; D020033:Protein Isoforms / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology; D012995:Solubility
D002352:Carrier Proteins; D005227:Fatty Acids; D047908:Intracellular Signaling Peptides and Proteins; D010940:Plant Proteins; D020033:Protein Isoforms; C087488:WAX9 protein, Brassica oleracea; C030363:lipid transfer protein; D019308:Palmitic Acid