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Biomedical subjects

K M Koeller

Publications and source records attributed to K M Koeller.

8 recordsLinked to original sources
PubMed2001

Enzymes for chemical synthesis.

New catalytic synthetic methods in organic chemistry that satisfy increasingly stringent environmental constraints are in great demand by the pharmaceutical and chemical industries. In addition, novel catalytic procedures are necessary to produce the emerging classes of organic compounds that are becoming the targets of molecular and biomedical research. Enzyme-catalysed chemical transformations are now widely recognized as practical alternatives to traditional (non-biological) organic synthesis, and as convenient solutions to certain intractable synthetic problems.

Carbon↗
PubMed2000

Chemoenzymatic synthesis of sialyl-trimeric-Lewis x.

The decasaccharide sialyl-trimeric-Lewis x is a component of glycoproteins and glycolipids that serve as E- and P-selectin ligands. The synthesis of this target structure was accomplished by utilizing a combination of chemical and enzymatic methods. Highlights of the chemical synthesis include minimal use of protecting groups and regioselective glycosylations to arrive at a linear tri-lactosamine structure. Glycosyltransferase-catalyzed reactions were then employed for the addition of the terminal sialic acid and branch-point fucose residues. Notably, fucosyltransferases V and VI showed different specificities for the sialyl-tri-lactosamine core structure.

Carbohydrate Conformation↗
PubMed2000

Chemoenzymatic synthesis and fluorescent visualization of cell-surface selectin-bound sialyl Lewis x derivatives.

Sialyl Lewis x (sLe(x)) derivatives conjugated to readily visualized molecular labels are useful chemical probes to study selectin-carbohydrate interactions. Localization of the selectins on the surface of leukocytes and activated endothelial cells can be detected through fluorescence of bound selectin ligands. Herein we present a short chemoenzymatic synthesis of a fluorescently labeled bivalent sLe(x) conjugate. The use of an amino-substituted monovalent sLe(x) to obtain fluorescent- and biotin-labeled sLe(x) derivatives is also described. The cell-staining utility of the fluorescent sLe(x) conjugates is demonstrated for a HUVEC cell line expressing E-selectin and for CHO-K1 cells expressing either L- or E-selectin.

Alanine↗
PubMed2000

Chemoenzymatic synthesis of PSGL-1 glycopeptides: sulfation on tyrosine affects glycosyltransferase-catalyzed synthesis of the O-glycan.

PSGL-1 is the primary glycoprotein ligand for P-selectin during the inflammatory response. Interestingly, the N-terminal sequence, containing both a site of tyrosine sulfation and an O-glycan, has been shown to bind to P-selectin with an affinity similar to full-length PSGL-1. To further characterize this system, the synthesis of glycopeptides from PSGL-1 was undertaken. The synthesis involved both solution- and solid-phase synthesis, as well as enzymatic transformations. During the synthesis, notable reactivity differences of the glycosyltransferases toward sulfated and unsulfated versions of the same glycopeptides were observed.

Amino Acid Sequence↗
PubMed2000

Emerging themes in medicinal glycoscience.

The recognition of complex carbohydrates and glycoconjugates as mediators of important biological processes has stimulated investigation into their therapeutic potential. New approaches for the simplification of glycoconjugate synthesis are overcoming the limitations of existing methods and providing a diverse array of these biomolecules. As the accessibility of glycoconjugates increases, carbohydrate-based constructs are becoming available for analysis as medicinal agents in a wide range of therapies.

Anti-Infective Agents↗
PubMed2000

Complex carbohydrate synthesis tools for glycobiologists: enzyme-based approach and programmable one-pot strategies.

The ultimate goal in complex carbohydrate synthesis is to develop synthetic tools which are simple and easily accessible to glycobiologists. This review will describe methods which have the potential to reach this goal, with particular focus on enzymatic and computer-based one-pot approaches for the preparation of complex carbohydrates and glycoconjugates.

Carbohydrate Sequence↗
PubMed1999

Expression of alpha2,8/2,9-polysialyltransferase from Escherichia coli K92. Characterization of the enzyme and its reaction products.

The capsular polysaccharide of Escherichia coli K92 contains alternating -8-NeuAcalpha2- and -9-NeuAcalpha2- linkages. The enzyme catalyzing this polymerizing reaction has been cloned from the genomic DNA of E. coli K92. The 1.2-kilobase polymerase chain reaction fragment was subcloned in pRSET vector and the protein was expressed in the BL21(DE3) strain of E. coli with a hexameric histidine at its N-terminal end. The enzyme was isolated in the supernatant after lysis of the cells and fractionated by ultracentrifugation. Western blotting using anti-histidine antibody showed the presence of a band that migrated at about 47.5 kDa on both reducing and nonreducing SDS-polyacrylamide gel electrophoresis, indicating a monomeric enzyme. Among the carbohydrate acceptors tested, N-acetylneuraminic acid and the gangliosides G(D3) and G(Q1b) were preferred substrates. The cell-free enzyme reaction products obtained were characterized by NMR and mass spectrometry, which indicated the presence of both alpha2,9- and alpha2,8-linked polysialyl structure. The K92 neuS gene was used to transform the K1 strain of E. coli, the capsule of which contains only -8-NeuAcalpha2- linkages. Analysis of the polysaccharides isolated from these transformed cells is consistent with the presence of both -8-NeuAcalpha2- and -9-NeuAcalpha2- linkages. Our results suggest that the neuS gene product of E. coli K92 catalyzes the synthesis of polysialic acid with alpha2,9- and alpha2,8-linkages in vitro and in vivo.

Blotting, Western↗