title: Purification, physico-chemical characterization and thermodynamics of chitooligosaccharide binding to cucumber (Cucumis sativus) phloem lectin.
abstract: A chitooligosaccharide-specific lectin has been purified from the phloem exudate of cucumber (Cucumis sativus) by affinity chromatography on chitin. The molecular weight of the cucumber phloem lectin (CPL) was determined as 51912.8Da by mass spectrometry whereas SDS-PAGE yielded a single band with a subunit mass of 26kDa, indicating that the protein is a homodimer. Peptide mass fingerprinting studies strongly suggest that CPL is identical to the 26kDa phloem protein 2 (PP2) from cucumber. CD spectroscopy indicated that CPL is a predominantly β-sheets protein. Hemagglutination activity of CPL was mostly unaffected between 4 and 90°C and between pH 4.0 and 10.0, indicating functional stability of the protein. Isothermal titration calorimetric studies indicate that the CPL dimer binds to two chitooligosaccharide ((GlcNAc)2-6) molecules with association constants ranging from 1.0×103to 17.5×105M-1. The binding reaction was strongly enthalpy driven (ΔHb=-ve) with negative contribution from binding entropy (ΔSb=-ve). The enthalpy-driven nature of binding reactions suggests that hydrogen bonding and van der Waals interactions stabilize the CPL-chitooligosaccharide association. Enthalpy-entropy compensation was observed for the CPL-chitooligosaccharide interaction, indicating that water molecules play an important role in the binding process.
authors: School of Chemistry, University of Hyderabad, Hyderabad 500046, India(1).
authors: Pavan Kumar
authors: PK
authors: Nareddy
authors: School of Chemistry, University of Hyderabad, Hyderabad 500046, India(1).
authors: Kishore Babu
authors: KB
authors: Bobbili
authors: School of Chemistry, University of Hyderabad, Hyderabad 500046, India(1). Electronic address: mjswamy1@gmail.com.
authors: Musti J
authors: MJ
authors: Swamy
mesh_terms: D000595:Amino Acid Sequence; D048271:Chitosan / Q000737:chemistry*; D018553:Cucumis sativus / Q000737:chemistry*; D019277:Entropy; D006863:Hydrogen-Ion Concentration; D009844:Oligosaccharides / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D037121:Plant Lectins / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D017433:Protein Structure, Secondary; D013696:Temperature; D013816:Thermodynamics
chemical_list: D009844:Oligosaccharides; D037121:Plant Lectins; C091028:phloem lectin; D048271:Chitosan
keywords: Cucurbitaceae; Isothermal titration calorimetry; PP2 protein