Caenorhabditis elegans proteins captured by immobilized Galβ1-4Fuc disaccharide units: assignment of 3 annexins

Abstract

Abbreviations: ACN, acetonitrile; EDTA, ethylenediaminetetraacetic acid; LC-MS/MS, liquid chromatography-tandem mass spectrometry; TFA, trifluoroacetic acid Pre-print version Takeuchi T. et al. Carbohydrate Research 346 (2011) Adsorbed C. elegans proteins were eluted with ethylenediaminetetraacetic acid (EDTA) and followed by lactose (Galβ1-4Glc), digested with trypsin, and were then subjected to proteomic analysis using LC-MS/MS. Three annexins, namely NEX-1, -2, and -3, were assigned in the EDTA-eluted fraction. Whereas, galectins, namely LEC-1, -2, -4, -6, -9, -10, and DC2.3a, were assigned in the lactose-eluted fraction. The affinity of annexins for Galβ1-4Fuc was further confirmed by adsorption of recombinant NEX-1, -2, and -3 proteins to the Galβ1-4Fuc column in the presence of Ca 2+ . Furthermore, frontal affinity chromatography analysis using an immobilized NEX-1 column showed that NEX-1 has an affinity for Galβ1-4Fuc, but no affinity towards Galβ1-3Fuc and Galβ1-4GlcNAc.We would hypothesize that the recognition of the Galβ1-4Fuc disaccharide unit is involved in some biological processes in C. elegans and other species of the Pre-print version Takeuchi T. et al.

The Effects of Levetiracetam on Cerebrospinal Fluid and Plasma NPY and GAL, and on the Components of Stress Response System, hs-CRP, and S100B Protein in Serum of Patients with Refractory Epilepsy

“…It is reported that NPY may be involved in the abnormal electrical activity in the vulnerable hippocampus neurons during epileptic seizure [38], and its levels increase in different types of epilepsy [39]. GAL is also widely distributed in the central and peripheral nervous systems and acts through G protein-coupled receptor that play an important role in the control of seizures in animal epilepsy [10] [40]. In this study, we showed that levetiracetam markedly lowered NPY and GAL levels in plasma and CSF in patients with refractory epilepsy (P \ 0.05).…”

Affinity Chromatography of Lectins

“…They also provide an eight carbon spacer arm. In carbohydrate affinity chromatography, the method is appropriate for coupling glycoproteins and carbohydrates or, more often, carbohydrate derivatives containing an amino group (Takeuchi et al, 2011).…”

“…A spacer group can be attached to the carrier (Section 4.2), or alternatively, sugar derivatives can be synthesized by attaching spacers, such as 6-amino-hexyl and ε-aminocaproyl groups (Lis & Sharon, 1981). Several carbohydrate derivatives have recently been synthesized and utilized in affinity chromatography for lectin isolations, such as the Galǐ1-4Fuc derivative containing a hydrophilic spacer modified with a free amino group (Takeuchi et al, 2011), an iodoacetamidyl derivative of Glc 1 Man 9 GlcNAc 2 (Watanabe et al, 2007), and a derivative of L-fucose with a BSA spacer (Argayosa & Lee, 2009). In addition, a novel method for covalent immobilization of a range of carbohydrate derivatives onto polymeric resin beads has been described .…”

“…The most widely used have been differently activated Sepharoses -divinyl sulphone (Almanza et al, 2004;Lavanya Latha et al, 2006;Pohleven et al, 2011) and epoxy (epichlorohydrin) activated Sepharose have been used to immobilize carbohydrates (Chen et al, 1999;Kaur et al, 2005;Maheswari et al, 2002;Mora et al, 2008;Nagata, 2005), and cyanogen bromide activated Sepharose for glycoproteins (Bhat et al, 2010;Bhowal et al, 2005;Kajiya et al, 2003;Mo et al, 1999;Naganuma et al, 2006;Nagre et al, 2010;Vega & Pérez, 2006;Yang et al, 2007). Nhydroxysuccinimide Sepharose was used for coupling with sugar derivatives containing amino groups (Takeuchi et al, 2011), and thiopropyl Sepharose for immobilizing iodoacetamidyl glycan derivatives (Watanabe et al, 2007). Mini-Leak agarose (Kem-En-Tec), a divinyl sulphone activated matrix (Valadez-Vega et al, 2011), and Seralose (Konozy et al, 2002) have also been used.…”

Purification of galectin-1 mutants using an immobilized Galactoseβ1–4Fucose affinity adsorbent

“…However, as evidenced by site-directed mutagenesis, each of the eight amino acid residues in LEC-6 had a relatively small impact on the interaction with Galb1-4Fuc compared to that with Galb1-4GlcNAc [16]. Furthermore, experiments with immobilized-Galb1-4Fuc have revealed that various C. elegans galectins have the affinity for Galb1-4Fuc disaccharide [20], although the eight amino acids are relatively less conserved in C. elegans galectins [21], for example, LEC-9, which has two substitutions among the 8 amino acid residues, could bind immobilized-Galb1-4Fuc. These results indicate that immobilized Galb1-4Fuc adsorbent may be useful for affinity purification of mutant galectins that cannot bind vertebrate-type glycans.…”