title: Gene/protein expression level, immunolocalization and binding characteristics of fatty acid binding protein from Clonorchis sinensis (CsFABP).
abstract: Clonorchis sinensis fatty acid-binding protein (CsFABP) belongs to a multigene family of lipid-binding proteins and is considered to be a promising vaccine candidate for human clonorchiasis. In this study, binding characteristics of CsFABP have been examined for the first time. The recombinant CsFABP (rCsFABP) was found to bind 11-(dansylamino) undecanoic acid (DAUDA), causing a blue shift in the fluorescence emission from 543 to 531 nm with an excitation wavelength of 345 nm and a substantial increase in fluorescence intensity. Fluorimetric titration of rCsFABP with DAUDA exhibited an apparent dissociation constant (K (d)) of 1.58 ± 0.14 μM. In the competitive experiment, the rCsFABP efficiently bound saturated C(10)-C(18) fatty acids and unsaturated fatty acids (oleic acid and linoleic acid), and the latter presented the higher affinity. Furthermore, quantitative RT-PCR and western blotting analysis revealed that CsFABP mRNA and protein were differentially expressed throughout the developmental cycle stages of the parasite, which occur in the definitive host (metacercariae, adult worms, and eggs). In addition, immunolocalization assay showed that CsFABP was localized on the vitelline gland, tegument, intestine, seminal vesicle, eggs in uterus, ovary, and testicle of C. sinensis adult worm, as well as on the vitelline gland of metacercaria. Intriguingly, the surface tissue of the bile duct where C. sinensis resided in the infected Sprague-Dawley rat was also strongly labeled, implying that CsFABP may possibly mediate direct interactions with host cells as a component of excretory/secretory products.
authors.affiliation: Department of Parasitology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou 510080, People's Republic of China.
authors.forename: Lisi
authors.initials: L
authors.lastname: Huang
authors.forename: Yue
authors.initials: Y
authors.lastname: Hu
authors.forename: Yan
authors.initials: Y
authors.lastname: Huang
authors.forename: Hang
authors.initials: H
authors.lastname: Fang
authors.forename: Ran
authors.initials: R
authors.lastname: Li
authors.forename: Dong
authors.initials: D
authors.lastname: Hu
authors.forename: Wenfang
authors.initials: W
authors.lastname: Li
authors.forename: Xuerong
authors.initials: X
authors.lastname: Li
authors.forename: Chi
authors.initials: C
authors.lastname: Liang
authors.forename: Xinbing
authors.initials: X
authors.lastname: Yu
mesh_terms: D000818:Animals; D001652:Bile Ducts / Q000378:metabolism / Q000469:parasitology; D001667:Binding, Competitive; D015153:Blotting, Western; D003003:Clonorchiasis / Q000378:metabolism / Q000469:parasitology; D003004:Clonorchis sinensis / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D003619:Dansyl Compounds / Q000378:metabolism; D004195:Disease Models, Animal; D050556:Fatty Acid-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005227:Fatty Acids / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D015801:Helminth Proteins / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D018510:RNA, Helminth / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000378:metabolism; D013050:Spectrometry, Fluorescence
chemical_list: D003619:Dansyl Compounds; D050556:Fatty Acid-Binding Proteins; D005227:Fatty Acids; D015801:Helminth Proteins; D018510:RNA, Helminth; D012333:RNA, Messenger; D011994:Recombinant Proteins; C054400:11-(dansylamino)undecanoic acid