3554-96-9Relevant articles and documents
Facile one-step syntheses of modified O-glycoprotein Galβ1-3GalNAc structures by transglycosylation employing three β-galactosidases from bovine testes, Xanthomonas manihotis, and Bacillus circulans
Kroeger, Lars,Thiem, Joachim
, p. 717 - 722 (2005)
Natural O-glycoproteins such as the Thomsen-Friedenreich antigen or gangliosides contain the motif Galβ1-3GalNAc as an important disaccharide with significant biologic activity. The arrangement of spatial functionalities in this structure are of particular interest with regard to the development of potential leads en route to pharmaceuticals. Therefore, it was desired to obtain access to a range of modified derivatives of the aforementioned motif paying particular attention to introducing specific deoxy functions instead of hydroxyl groups. Copyright Taylor & Francis, Inc.
Fluorescence activated cell sorting as a general ultra-high-throughput screening method for directed evolution of glycosyltransferases
Yang, Guangyu,Rich, Jamie R.,Gilbert, Michel,Wakarchuk, Warren W.,Feng, Yan,Withers, Stephen G.
experimental part, p. 10570 - 10577 (2010/09/17)
Glycosyltransferases (GTs) offer very attractive approaches to the synthesis of complex oligosaccharides. However, the limited number of available GTs, together with their instability and strict substrate specificity, have severely hampered the broad application of these enzymes. Previous attempts to broaden the range of substrate scope and to increase the activity of GTs via protein engineering have met with limited success, partially because of the lack of effective high-throughput screening methods. Recently, we reported an ultra-high-throughput screening method for sialyltransferases based on fluorescence-activated cell sorting (Aharoni et al. Nat. Methods 2006, 3, 609-614). Here, we considerably improve this method via the introduction of a two-color screening protocol to minimize the probability of false positive mutants and demonstrate its generality through directed evolution of a neutral sugar transferase, β-1,3-galactosyltransferase CgtB. A variant with broader substrate tolerance than the wild-type enzyme and 300-fold higher activity was identified rapidly from a library of >107 CgtB mutants. Importantly, the variant effected much more efficient synthesis of G M1a and asialo GM1 oligosaccharides, the building blocks of important therapeutic glycosphingolipids, than did the parent enzyme. This work not only establishes a new methodology for the directed evolution of galactosyltransferases, but also suggests a powerful strategy for the screening of almost all GT activities, thereby facilitating the engineering of glycosyltransferases.
Non-crosslinked protein particles for therapeutic and diagnostic use
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, (2008/06/13)
Albumin particles in the nanometer and micrometer size range in an aqueous suspension are rendered stable against resolubilization without the aid of a crosslinking agent and without denaturation, by the incorporation of a stabilizing agent in the particle composition. Particles which are primarily albumin in the nanometer and micrometer size range in an aqueous suspension are rendered stable against resolublization by the incorporation of a reducing agent, oxidizing agent, hydrogen-accepting molecule, high molecular weight polymer, sulfur-containing ring compound or combinations thereof.
