ABSTRACT
Tetanus toxoid protein has been characterized with regard oligomeric state and hydrodynamic (low-resolution) shape, important parameters with regard its use in glycoconjugate vaccines. From sedimentation velocity and sedimentation equilibrium analysis in the analytical ultracentrifuge tetanus toxoid protein is shown to be mostly monomeric in solution (~86%) with approximately 14% dimer. The relative proportions do not appear to change significantly with concentration, suggesting the two components are not in reversible equilibrium. Hydrodynamic solution conformation studies based on high precision viscometry, combined with sedimentation data show the protein to be slightly extended conformation in solution with an aspect ratio ~3. The asymmetric structure presents a greater surface area for conjugation with polysaccharide than a more globular structure, underpinning its popular choice as a conjugation protein for glycoconjugate vaccines.
Subject(s)
Glycoconjugates , Tetanus Toxoid/chemistry , Vaccines, Conjugate/chemistry , Chromatography, Gel , Molecular Weight , Protein Multimerization , Scattering, Small Angle , Ultracentrifugation , ViscosityABSTRACT
Capsular polysaccharides from ten different serotypes of Streptococcus pneumoniae have been studied with regards their hydrodynamic properties in solution, namely their sedimentation coefficient and molar mass distributions, solution conformations and flexibilities (persistence lengths Lp), important properties for the construction of polysaccharide and glycoconjugate vaccines. Sedimentation and molar mass distributions (obtained by sedimentation velocity and equilibrium analysis in the analytical ultracentrifuge supported by size exclusion chromatography coupled to multi-angle light scattering measurements) were generally unimodal, with weight (mass) average molar masses ranging from 100×10(3) to 1300×10(3) g/mol. Estimates of chain flexibilities from three different procedures applied to intrinsic viscosity, sedimentation coefficient and molar mass data, showed that the polysaccharides from all the serotypes studied had semi-flexible structures in solution with persistence lengths in the range from â¼4 to 9 nm.
Subject(s)
Polysaccharides, Bacterial/chemistry , Polysaccharides, Bacterial/isolation & purification , Streptococcus pneumoniae/chemistry , Streptococcus pneumoniae/isolation & purification , Solubility , Ultracentrifugation/methodsABSTRACT
A novel series of tertiary alcohol containing 2-substituted benzyl morpholines have been discovered as potent and selective inhibitors of the norepinephrine transporter. Efficient synthetic routes were developed featuring a highly diastereoselective nucleophilic addition of benzyl Grignard reagents to enantiopure (4-benzylmorpholin-2-yl)phenylmethanone (11) as the key synthetic step. In vitro binding affinity for the norepinephrine, dopamine and serotonin transporters and in vivo examination of a select compound (16) in a pharmacodynamic animal model for norepinephrine reuptake inhibition are presented.
