Paul Kosma

Publications of Paul Kosma

• Program and abstracts for the 2011 Meeting of the Society for Glycobiology.

  Authors: M Tony Hollingsworth, Gerald W Hart, James C Paulson, Elizabeth Stansell, Kevin Canis, I-Cheuh Huang, Maria Panico, Howard Morris, Stuart Haslam, Michael Farzan [......] Kazunori Hamamura, Takenosuke Yoshida, Kaoru Akita, Tetsuya Okajima, Keiko Furukawa, Takeshi Urano, Koichi Furukawa, L Renee Ruhaak, Suzanne Miyamoto, Carlito B Lebrilla

  Glycobiology. 11/2011; 21(11):1454-531.

  Cell surface mucins configure the cell surface by presenting extended protein backbones that are heavily O-glycosylated. The glycopeptide structures establish physicochemical properties at the cellCell surface mucins configure the cell surface by presenting extended protein backbones that are heavily O-glycosylated. The glycopeptide structures establish physicochemical properties at the cell surface that enable and block the formation of biologically important molecular complexes. Some mucins, such as MUC1, associate with receptor tyrosine kinases and other cell surface receptors, and engage in signal transduction in order to communicate information regarding conditions at the cell surface to the nucleus. In that context, the MUC1 cytoplasmic tail (MUC1CT) receives phosphorylation signals from receptor tyrosine kinases and serine/threonine kinases, which enables its association with different signaling complexes that conduct these signals to the nucleus and perhaps other subcellular organelles. We have detected the MUC1CT at promoters of over 500 genes, in association with several different transcription factors, and have shown that promoter occupancy can vary under different growth factor conditions. However, the full biochemical nature of the nuclear forms of MUC1 and its function at these promoter regions remain undefined. I will present evidence that nuclear forms of the MUC1CT include extracellular and cytoplasmic tail domains. In addition, I will discuss evidence for a hypothesis that the MUC1CT possesses a novel catalytic function that enables remodeling of the transcription factor occupancy of promoters, and thereby engages in regulation of gene expression.

• Burkholderia cenocepacia BC2L-C is a super lectin with dual specificity and proinflammatory activity.

  Authors: Ondřej Sulák, Gianluca Cioci, Emilie Lameignère, Viviane Balloy, Adam Round, Irina Gutsche, Lenka Malinovská, Michel Chignard, Paul Kosma, Daniel F Aubert, Cristina L Marolda, Miguel A Valvano, Michaela Wimmerová, Anne Imberty

  PLoS pathogens. 09/2011; 7(9):e1002238.

  Lectins and adhesins are involved in bacterial adhesion to host tissues and mucus during early steps of infection. We report the characterization of BC2L-C, a soluble lectin from the opportunisticLectins and adhesins are involved in bacterial adhesion to host tissues and mucus during early steps of infection. We report the characterization of BC2L-C, a soluble lectin from the opportunistic pathogen Burkholderia cenocepacia, which has two distinct domains with unique specificities and biological activities. The N-terminal domain is a novel TNF-α-like fucose-binding lectin, while the C-terminal part is similar to a superfamily of calcium-dependent bacterial lectins. The C-terminal domain displays specificity for mannose and l-glycero-d-manno-heptose. BC2L-C is therefore a superlectin that binds independently to mannose/heptose glycoconjugates and fucosylated human histo-blood group epitopes. The apo form of the C-terminal domain crystallized as a dimer, and calcium and mannose could be docked in the binding site. The whole lectin is hexameric and the overall structure, determined by electron microscopy and small angle X-ray scattering, reveals a flexible arrangement of three mannose/heptose-specific dimers flanked by two fucose-specific TNF-α-like trimers. We propose that BC2L-C binds to the bacterial surface in a mannose/heptose-dependent manner via the C-terminal domain. The TNF-α-like domain triggers IL-8 production in cultured airway epithelial cells in a carbohydrate-independent manner, and is therefore proposed to play a role in the dysregulated proinflammatory response observed in B. cenocepacia lung infections. The unique architecture of this newly recognized superlectin correlates with multiple functions including bacterial cell cross-linking, adhesion to human epithelia, and stimulation of inflammation.

• Crystal and molecular structure of methyl L-glycero-α-D-manno-heptopyranoside, and synthesis of 1→7 linked L-glycero-D-manno-heptobiose and its methyl α-glycoside.

  Authors: Daniel Artner, Christian Stanetty, Kurt Mereiter, Alla Zamyatina, Paul Kosma

  Carbohydrate research. 06/2011; 346(13):1739-46.

  Methyl l-glycero-α-d-manno-heptopyranoside was synthesized in good yield by a Fischer-type glycosylation of the heptopyranose with methanol in the presence of cation-exchange resin under reflux andMethyl l-glycero-α-d-manno-heptopyranoside was synthesized in good yield by a Fischer-type glycosylation of the heptopyranose with methanol in the presence of cation-exchange resin under reflux and microwave conditions, respectively. The compound crystallized from 2-propanol in an orthorhombic lattice of space group P2(1)2(1)2 showing a comparatively porous structure with a 2-dimensional O-H⋯O hydrogen bond network. As model compounds for the side chain domains of the inner core structure of bacterial lipopolysaccharide, l-glycero-α-d-manno-heptopyranosyl-(1→7)-l-glycero-d-manno-heptopyranose and the corresponding disaccharide methyl α-glycoside were prepared. The former compound was generated via glycosylation of a benzyl 5,6-dideoxy-hept-5-enofuranoside intermediate followed by catalytic osmylation and deprotection. The latter disaccharide was efficiently synthesized in good yield by a straightforward coupling of an acetylated N-phenyltrifluoroacetimidate heptopyranosyl donor to a methyl 2,3,4,6-tetra-O-acetyl heptopyranoside acceptor derivative followed by Zemplén deacetylation.

• Structural insights into parallel strategies for germline antibody recognition of lipopolysaccharide from Chlamydia.

  Authors: Dylan W Evans, Sven Müller-Loennies, Cory L Brooks, Lore Brade, Paul Kosma, Helmut Brade, Stephen V Evans

  Glycobiology. 05/2011; 21(8):1049-59.

  The structure of the antigen-binding fragment from the monoclonal antibody S64-4 in complex with a pentasaccharide bisphosphate fragment from chlamydial lipopolysaccharide has been determined byThe structure of the antigen-binding fragment from the monoclonal antibody S64-4 in complex with a pentasaccharide bisphosphate fragment from chlamydial lipopolysaccharide has been determined by x-ray diffraction to 2.6 Å resolution. Like the well-characterized antibody S25-2, S64-4 displays a pocket formed by the residues of germline sequence corresponding to the heavy and light chain V gene segments that binds the terminal Kdo residue of the antigen; however, although S64-4 shares the same heavy chain V gene segment as S25-2, it has a different light chain V gene segment. The new light chain V gene segment codes for a combining site that displays greater affinity, different specificity, and allows a novel antigen conformation that brings a greater number of antigen residues into the combining site than possible in S25-2. Further, while antibodies in the S25-2 family use complementarity determining region (CDR) H3 to discriminate among antigens, S64-4 achieves its specificity via the new light chain V gene segment and resulting change in antigen conformation. These structures reveal an intriguing parallel strategy where two different combinations of germline-coded V gene segments can act as starting points for the generation of germline antibodies against chlamydial antigens and show how anti-carbohydrate antibodies can exploit the conformational flexibility of this class of antigens to achieve high affinity and specificity independently of CDR H3.

• Synthesis of new glycyrrhetinic acid derived ring A azepanone, 29-urea and 29-hydroxamic acid derivatives as selective 11β-hydroxysteroid dehydrogenase 2 inhibitors.

  Authors: Rawindra Gaware, Rupesh Khunt, Laszlo Czollner, Christian Stanetty, Thierry Da Cunha, Denise V Kratschmar, Alex Odermatt, Paul Kosma, Ulrich Jordis, Dirk Classen-Houben

  Bioorganic & medicinal chemistry. 03/2011; 19(6):1866-80.

  Glycyrrhetinic acid, the metabolite of the natural product glycyrrhizin, is a well known nonselective inhibitor of 11β-hydroxysteroid dehydrogenase (11β-HSD) type 1 and type 2. Whereas inhibition ofGlycyrrhetinic acid, the metabolite of the natural product glycyrrhizin, is a well known nonselective inhibitor of 11β-hydroxysteroid dehydrogenase (11β-HSD) type 1 and type 2. Whereas inhibition of 11β-HSD1 is currently under consideration for treatment of metabolic diseases, such as obesity and diabetes, 11β-HSD2 inhibitors may find therapeutic applications in chronic inflammatory diseases and certain forms of cancer. Recently, we published a series of hydroxamic acid derivatives of glycyrrhetinic acid showing high selectivity for 11β-HSD2. The most potent and selective compound is active against human 11β-HSD2 in the low nanomolar range with a 350-fold selectivity over human 11β-HSD1. Starting from the lead compounds glycyrrhetinic acid and the hydroxamic acid derivatives, novel triterpene type derivatives were synthesized and analyzed for their biological activity against overexpressed human 11β-HSD1 and 11β-HSD2 in cell lysates. Here we describe novel 29-urea- and 29-hydroxamic acid derivatives of glycyrrhetinic acid as well as derivatives with the Beckman rearrangement of the 3-oxime to a seven-membered ring, and the rearrangement of the C-ring from 11-keto-12-ene to 12-keto-9(11)-ene. The combination of modifications on different positions led to compounds comprising further improved selective inhibition of 11β-HSD2 in the lower nanomolar range with up to 3600-fold selectivity.

• A common NH53K mutation in the combining site of antibodies raised against chlamydial LPS glycoconjugates significantly increases avidity.

  Authors: Ryan J Blackler, Sven Müller-Loennies, Cory L Brooks, Dylan W Evans, Lore Brade, Paul Kosma, Helmut Brade, Stephen V Evans

  Biochemistry. 03/2011; 50(16):3357-68.

  The crystal structures of the antigen-binding fragment of the murine monoclonal antibody (mAb) S25-39 in the presence of several antigens representing chlamydial lipopolysaccharide (LPS) epitopesThe crystal structures of the antigen-binding fragment of the murine monoclonal antibody (mAb) S25-39 in the presence of several antigens representing chlamydial lipopolysaccharide (LPS) epitopes based on the bacterial sugar 3-deoxy-α-D-manno-oct-2-ulosonic acid (Kdo) have been determined at resolutions from 2.4 to 1.8 Å. The antigen-binding site of this antibody differs from the well-characterized antibody S25-2 by a single mutation away from the germline of asparagine H53 to lysine, yet this one mutation results in a significant increase in avidity across a range of antigens. A comparison of the two antibody structures reveals that the mutated Lys H53 forms additional hydrogen bonds and/or charged-residue interactions with the second Kdo residue of every antigen having two or more carbohydrate residues. Significantly, the NH53K mutation results from a single nucleotide substitution in the germline sequence common among a panel of antibodies raised against glycoconjugates containing carbohydrate epitopes of chlamydial LPS. Like S25-2, S25-39 displays significant induced fit of complementarity determining region (CDR) H3 upon antigen binding, with the unliganded structure possessing a conformation distinct from those reported earlier for S25-2. The four different observed conformations for CDR H3 suggest that this CDR has evolved to exploit the recognition potential of a flexible loop while minimizing the associated entropic penalties of binding by adopting a limited number of ordered conformations in the unliganded state. These observations reveal strategies evolved to balance adaptability and specificity in the germline antibody response to carbohydrate antigens.

• Synthesis of novel 3-amino and 29-hydroxamic acid derivatives of glycyrrhetinic acid as selective 11β-hydroxysteroid dehydrogenase 2 inhibitors.

  Authors: Christian Stanetty, Laszlo Czollner, Iris Koller, Priti Shah, Rawindra Gaware, Thierry Da Cunha, Alex Odermatt, Ulrich Jordis, Paul Kosma, Dirk Classen-Houben

  Bioorganic & medicinal chemistry. 11/2010; 18(21):7522-41.

  Glycyrrhetinic acid, the metabolite of the natural product glycyrrhizin, is a well known nonselective inhibitor of 11β-hydroxysteroid dehydrogenase (11β-HSD) type 1 and type 2. Whereas inhibition ofGlycyrrhetinic acid, the metabolite of the natural product glycyrrhizin, is a well known nonselective inhibitor of 11β-hydroxysteroid dehydrogenase (11β-HSD) type 1 and type 2. Whereas inhibition of 11β-HSD1 is currently under consideration for treatment of metabolic diseases, such as obesity and diabetes, 11β-HSD2 inhibitors may find therapeutic applications in chronic inflammatory diseases and certain forms of cancer. So far, no selective 11β-HSD2 inhibitor has been developed and neither animal studies nor clinical trials have been reported based on 11β-HSD2 inhibition. Starting from the lead compound glycyrrhetinic acid, novel triterpene type derivatives were synthesized and analyzed for their biological activity against overexpressed human 11β-HSD1 and 11β-HSD2 in cell lysates. Several hydroxamic acid derivatives showed high selectivity for 11β-HSD2. The most potent and selective compound is active against human 11β-HSD2 in the low nanomolar range with a 350-fold selectivity over human 11β-HSD1.

• Efficient Synthesis of 4-Amino-4-deoxy-L-arabinose and Spacer-equipped 4-Amino-4-deoxy-L-arabinopyranosides by Transglycosylation Reactions.

  Authors: Bernhard Müller, Markus Blaukopf, Andreas Hofinger, Alla Zamyatina, Helmut Brade, Paul Kosma

  Synthesis. 09/2010; 2010(18):3143-3151.

  Methyl 4-azido-4-deoxy-β-L-arabinopyranoside has been synthesized in five steps starting from methyl β-D-xylopyranoside in a multigram scale without chromatographic purification in 78% overall yield.Methyl 4-azido-4-deoxy-β-L-arabinopyranoside has been synthesized in five steps starting from methyl β-D-xylopyranoside in a multigram scale without chromatographic purification in 78% overall yield. The transformation relied on selective tosylation/nosylation at O-4 followed by acylation, S(N)2 displacement with sodium azide and subsequent deprotection. The methyl 4-azido-4-deoxy-arabinoside was then converted into allyl, propenyl, ω-bromohexyl and chlorethoxyethyl spacer glycosides by transglycosylation with the respective alcohols in good yields and fair anomeric selectivity. Reduction of the azido group and further transformations of the aglycon afforded ω-thiol-containing spacer derivatives. Coupling to maleimide-activated BSA provided a potent immunogen which was used to generate murine and rabbit polyclonal sera binding to LPS-core epitopes containing 4-amino-4-deoxy-arabinose residues.

• Foreword.

  Authors: Paul Kosma, Todd Lowary

  Carbohydrate research. 07/2010; 345(10):1251.

• Synthesis of a neoglycoconjugate containing a Chlamydophila psittaci-specific branched Kdo trisaccharide epitope.

  Authors: Paul Kosma, Andreas Hofinger, Sven Müller-Loennies, Helmut Brade

  Carbohydrate research. 03/2010; 345(5):704-8.

  The branched Kdo trisaccharide sodium (3-deoxy-alpha-D-manno-oct-2-ulopyranosyl)onate-(2-->8)-[sodium (3-deoxy-alpha-D-manno-oct-2-ulopyranosyl)onate-(2-->4)]-sodium (allylThe branched Kdo trisaccharide sodium (3-deoxy-alpha-D-manno-oct-2-ulopyranosyl)onate-(2-->8)-[sodium (3-deoxy-alpha-D-manno-oct-2-ulopyranosyl)onate-(2-->4)]-sodium (allyl 3-deoxy-alpha-D-manno-oct-2-ulopyranosid)onate has been prepared utilizing the regioselective glycosylation of the C-7, C-8 diol entity of a Kdo monosaccharide acceptor with a Kdo bromide donor followed by the attachment of the third Kdo unit to O-4 of the disaccharide intermediate. Deacetylation and hydrolysis of the methyl ester groups furnished the trisaccharide allyl glycoside which was converted into the corresponding 3-(2-aminoethylthio)propyl glycoside. Subsequent covalent attachment to bovine serum albumin furnished a neoglycoconjugate serving as an antigen for the induction of Chlamydophila psittaci-specific monoclonal antibodies.

• Analysis of Cross-Reactive and Specific Anti-Carbohydrate Antibodies against Lipopolysaccharide from Chlamydophila psittaci.

  Authors: Sandra Gerstenbruch, Cory L Brooks, Paul Kosma, Lore Brade, C Roger Mackenzie, Stephen V Evans, Helmut Brade, Sven Müller-Loennies

  Glycobiology. 12/2009;

  Chlamydiae contain a rough type lipopolysaccharide (LPS) of 3-deoxy-alpha-D-manno-oct-2-ulopyranosonic acid residues (Kdo). Two Kdo trisaccharides, 2.8/2.4- and 2.4/2.4-linked, and a branchedChlamydiae contain a rough type lipopolysaccharide (LPS) of 3-deoxy-alpha-D-manno-oct-2-ulopyranosonic acid residues (Kdo). Two Kdo trisaccharides, 2.8/2.4- and 2.4/2.4-linked, and a branched 2.4[2.8]2.4-linked Kdo tetrasaccharide occur in Chlamydiaceae. While the 2.8/2.4-linked trisaccharide contains a family-specific epitope, the branched Kdo oligosaccharide occurs only in Chlamydophila psittaci and antibodies against it will be useful in human and veterinarian diagnostics. To overcome the generation of cross-reactive antibodies that bind with high affinity to a dominant epitope formed by 2.4/2.4-linked Kdo, we immunized mice with a synthetic 2.4[2.8]-linked branched Kdo trisaccharide and used phage display of scFv to isolate recombinant antibody fragments (NH2240-31 and SAG506-01) that recognize the branched Kdo oligosaccharide with a K(D) of less than 10 nM. Importantly, although these antibodies used germline genes coding for an inherited Kdo recognition site they were able clearly to distinguish between 2.4[2.8]2.4- and 2.4/2.4-linked Kdo. Sequence determination, binding data, and X-ray structural analysis revealed the basis for the improved discrimination between similar Kdo ligands and indicated that the alteration of a stacking interaction from a phenylalanine residue in the center of the combining site to a tyrosine residue facing away from the center favours recognition of branched 2.4[2.8]2.4-linked Kdo residues. Immunofluorescence tests of infected cell monolayers using this antibody show specific staining of C. psittaci elementary bodies that allow it to be distinguished from other pathogenic chlamydiae.

• Monoclonal antibody S60-4-14 reveals diagnostic potential in the identification of Pseudomonas aeruginosa in lung tissues of cystic fibrosis patients.

  Authors: Kai Schmengler, Torsten Goldmann, Lore Brade, Patricia M Sánchez Carballo, Sandra Albrecht, Helmut Brade, Paul Kosma, Hany Sahly, Hans-Peter Hauber, Florian Länger, Rudolf Meyer, Tobias Welte, Ulrich Zähringer

  European journal of cell biology. 12/2009;

  The lipopolysaccharide (LPS) of Pseudomonas aeruginosa has been identified to contain an inner-core structure expressing a Pseudomonas-specific epitope. This target structure is characterized by aThe lipopolysaccharide (LPS) of Pseudomonas aeruginosa has been identified to contain an inner-core structure expressing a Pseudomonas-specific epitope. This target structure is characterized by a highly phosphorylated and 7-O-carbamoyl-l-glycero-alpha-d-manno-heptopyranose (CmHep) and was found to be present in all human-pathogenic Pseudomonas species of the Palleroni (RNA)-classification I scheme. We raised and selected the monoclonal antibody S60-4-14 (mAb S60-4-14, subtype IgG1) from mice immunized with heat-killed Pseudomonas bacteria. The epitope of this mAb was found to reside in the inner-core structure of P. aeruginosa and, hence, successfully evaluated for the immunohistochemical detection of P. aeruginosa in formalin- or HOPE-fixed (Hepes-glutamic acid buffer-mediated organic solvent protection effect) and paraffin-embedded human lung tissue slices. Lung specimens, mainly from explanted lungs of cystic fibrosis (CF) patients, as well as P. aeruginosa isolates from patients suffering from CF and patients with extrapulmonar Pseudomonas infections were investigated by PCR, immunohistochemistry, and Western blot analysis with mAb S60-4-14. The results revealed an unequivocal coincidence of PCR and immunohistochemistry. Together with the Western blot results mAb S60-4-14 displays a potential diagnostic tool for the specific identification of P. aeruginosa in infected lungs of CF.

• Antibodies Raised Against Chlamydial Lipopolysaccharide Antigens Reveal Convergence in Germline Gene Usage and Differential Epitope Recognition.

  Authors: Cory L Brooks, Sven Müller-Loennies, Svetlana N Borisova, Lore Brade, Paul Kosma, Tomoko Hirama, C Roger MacKenzie, Helmut Brade, Stephen V Evans

  Biochemistry. 12/2009;

  The structures of antigen-binding fragments from two related monoclonal antibodies have been determined to high resolution in the presence of several carbohydrate antigens raised against chlamydialThe structures of antigen-binding fragments from two related monoclonal antibodies have been determined to high resolution in the presence of several carbohydrate antigens raised against chlamydial lipopolysaccharide. With the exception of CDR H3, antibodies S54-10 and S73-2 are both derived from the same set of germline gene segments as the previously reported structures S25-2 and S45-18. Despite this similarity, the antibodies differ in specificity and the mechanism by which they recognize their cognate antigen. S54-10 uses an unrelated CDR H3 to recognize its antigen in a fashion analogous to S45-18; however, S73-2 recognizes the same antigen as S45-18 and S54-10 in a wholly unrelated manner. Together, these antibody-antigen structures provide snapshots into how the immune system uses the same set of inherited germline gene segments to generate multiple possible specificities that allow for differential recognition of epitopes, and how unrelated CDR H3 sequences can result in convergent binding of clinically-relevant bacterial antigens.

• Synthesis of glycyrrhetinic acid derivatives for the treatment of metabolic diseases.

  Authors: Igor Beseda, Laszlo Czollner, Priti S Shah, Rupesh Khunt, Rawindra Gaware, Paul Kosma, Christian Stanetty, Maria Carmen Del Ruiz Ruiz, Hassan Amer, Kurt Mereiter, Thierry Da Cunha, Alex Odermatt, Dirk Claßen-Houben, Ulrich Jordis

  Bioorganic & medicinal chemistry. 10/2009;

  The effect of glycyrrhetinic acid (GA) and GA-derivatives towards 11beta-hydroxysteroid dehydrogenase (11beta-HSD) was investigated. Novel compounds with modifications at positions C-3, C-11 and C-29The effect of glycyrrhetinic acid (GA) and GA-derivatives towards 11beta-hydroxysteroid dehydrogenase (11beta-HSD) was investigated. Novel compounds with modifications at positions C-3, C-11 and C-29 of the GA skeleton were prepared. Single crystal X-ray diffraction data of selected substances are reported and discussed.

• The role of CDR H3 in Antibody Recognition of a Synthetic Analogue of a Lipopolysaccharide Antigen.

  Authors: Cory L Brooks, Ryan J Blackler, Georg Sixta, Paul Kosma, Sven Müller-Loennies, Lore Brade, Tomoko Hirama, C Roger MacKenzie, Helmut Brade, Stephen V Evans

  Glycobiology. 09/2009;

  In order to explore the structural basis for adaptability in near germline monoclonal antibodies (mAb), we have examined the specificity of the promiscuous mAb S67-27 to both naturally derivedIn order to explore the structural basis for adaptability in near germline monoclonal antibodies (mAb), we have examined the specificity of the promiscuous mAb S67-27 to both naturally derived carbohydrate antigens and a variety of synthetic non-natural antigens based on the bacterial lipopolysaccharide component 3-deoxy-alpha-d-manno-oct-2-ulosonic acid (Kdo). One such analogue, a 7-O-methyl (7-O-Me) Kdo disaccharide, was found to bind to the antibody with at least 30 fold higher affinity than any other antigen tested. The structure of S67-27 in complex with this analogue and three other naturally occurring Kdo antigens revealed that the enhanced affinity of the mAb for the synthetic analogue was accomplished by the strategic positioning of CDR H3 away from a conserved Kdo binding pocket that allowed the formation of new antibody-antigen contacts. Furthermore, comparison of this structure with the structures of related mAbs revealed how the position and structure of CDR H3 influence the specificity or promiscuity of near-germline carbohydrate-recognizing antibodies by altering the architecture of the combining site.

• Synthesis and antigenic properties of C-7-modified Kdo mono- and disaccharide ligands and Kdo disaccharide interresidue lactones.

  Authors: Georg Sixta, Kurt Wimmer, Andreas Hofinger, Helmut Brade, Paul Kosma

  Carbohydrate research. 07/2009;

  In order to define binding interactions of Kdo-specific monoclonal antibodies directed against the chlamydial alpha-(2-->8)-linked Kdo disaccharide epitope on a molecular level, modifications at theIn order to define binding interactions of Kdo-specific monoclonal antibodies directed against the chlamydial alpha-(2-->8)-linked Kdo disaccharide epitope on a molecular level, modifications at the 7-position of the proximal and distal Kdo unit were investigated. The synthesis of 7-O-methyl and 7-azido-7-deoxy-7-epi-Kdo monosaccharide derivatives was achieved via an 8-O-TBS protected derivative, whereas methylation of O-7 at the proximal Kdo unit of the alpha-(2-->8)-linked Kdo disaccharide was conveniently accomplished via a 4,5; 4',5'; 7',8'-tri-O-carbonyl-protected disaccharide intermediate. Attempted epimerization at C-5 of the inner unit of a alpha-(2-->4)-linked Kdo disaccharide, however, resulted in formation of the corresponding 5,6-dehydro derivative, which was fully deprotected. Treatment of unprotected alpha-(2-->8)- as well as alpha-(2-->4)-linked Kdo disaccharides in neat acetic acid furnished the corresponding interresidue lactone derivatives. The lactones displayed limited stability under neutral conditions and were hydrolyzed at pH 7 within 3days. Access to the lactones, however, provides a means for selective derivatization of the carboxylic group located at the distal Kdo residue, which was demonstrated by methanolysis of the lactone to afford the monomethyl ester of the alpha-(2-->8)-linked Kdo disaccharide. ELISA inhibition experiments of the ligands with two Kdo-specific monoclonal antibodies showed slightly reduced reactivity for the binding of the alpha-(2-->8) Kdo-specific antibody S25-2, whereas the 7-O-methyl disaccharide antigen displayed high binding affinity toward the Kdo monosaccharide-specific antibody S67-27.

• Efficient synthesis of glycyrrhetinic acid glycoside/glucuronide derivatives using silver zeolite as promoter.

  Authors: Maria Carmen Del Ruiz Ruiz, Hassan Amer, Christian Stanetty, Igor Beseda, Laszlo Czollner, Priti Shah, Ulrich Jordis, Bernhard Kueenburg, Dirk Claßen-Houben, Andreas Hofinger, Paul Kosma

  Carbohydrate research. 05/2009;

  3-O-Glycopyranosides of glycyrrhetinic acid have been synthesized in good to high yields and excellent stereoselectivity using glycosyl bromide donors and silver zeolite as promoter. In addition to3-O-Glycopyranosides of glycyrrhetinic acid have been synthesized in good to high yields and excellent stereoselectivity using glycosyl bromide donors and silver zeolite as promoter. In addition to the preparation of glycosides containing beta-linked glucosyl, 2-deoxy-2-trichloroacetamido-glucosyl, galactosyl, cellobiosyl and lactosyl residues, also the deactivated acetylated methyl glucopyranosyluronate bromide donor could be coupled to triterpene aglycon ester derivatives in good yields. The ester protecting group located at C-30 of the oleanolic acid scaffold exerted an influence on the overall yield, with the methylester-protected glycosyl acceptor giving better yields compared to the allyl, benzyl as well as diphenylmethyl ester aglycon. The acetyl-protected glucuronides were differently deblocked in high yields via Zemplén deacetylation or via hydrogenolysis followed by Zemplén deacetylation, and alkaline hydrolysis, respectively, to allow for a selective liberation of the ester groups from either the glucuronide or the glycyrrhetinic acid unit, respectively. The target glycosides/glucuronides serve as probes for pharmaceutical studies aimed at defining structure-activity relationships of glycoside/glucuronide triterpenes.

• Pseudo-symmetry and twinning in crystals of homologous antibody Fv fragments.

  Authors: Cory L Brooks, Ryan J Blackler, Sandra Gerstenbruch, Paul Kosma, Sven Müller-Loennies, Helmut Brade, Stephen V Evans

  Acta crystallographica. Section D, Biological crystallography. 01/2009; 64(Pt 12):1250-8.

  A difference of seven conservative amino-acid substitutions between two single-chain antibodies (scFvs) specific for chlamydial lipopolysaccharide does not significantly affect their molecularA difference of seven conservative amino-acid substitutions between two single-chain antibodies (scFvs) specific for chlamydial lipopolysaccharide does not significantly affect their molecular structures or packing contacts, but dramatically affects their crystallization. The structure of the variable domain (Fv) of SAG173-04 was solved to 1.86 A resolution and an R(cryst) of 18.9% in space group P2(1)2(1)2(1). Crystals of the homologous SAG506-01 diffracted to 1.95 A resolution and appeared at first to have Patterson symmetry I4/m or P4/mmm; however, no solution could be found in space groups belonging to the former and refinement in the only solution corresponding to the latter (in space group P4(3)2(1)2) stalled at R(free) = 30.0%. Detailed examination of the diffraction data revealed that the crystal was likely to be twinned and that the correct space group was P2(1)2(1)2(1). Both translational pseudo-symmetry and pseudo-merohedral twinning were observed in one crystal of SAG506-01 and pseudo-merohedral twinning was observed for a second crystal. The final R factor for SAG506-01 after refinement in P2(1)2(1)2(1) was 20.5%.

• Van der Waals versus hydrogen-bonding forces in a crystalline analog of cellotetraose: cyclohexyl 4'-O-cyclohexyl beta-D-cellobioside cyclohexane solvate.

  Authors: Yuko Yoneda, Kurt Mereiter, Christian Jaeger, Lothar Brecker, Paul Kosma, Thomas Rosenau, Alfred French

  Journal of the American Chemical Society. 01/2009; 130(49):16678-90.

  Hydrogen bonding is important in cellulosic and other carbohydrate structures, but the role of interactions between nonpolar groups is less understood. Therefore, we synthesized cyclohexyl 4'-OHydrogen bonding is important in cellulosic and other carbohydrate structures, but the role of interactions between nonpolar groups is less understood. Therefore, we synthesized cyclohexyl 4'-O cyclohexyl beta-D-cellobioside (8), a molecule that has two glucose rings and two nonpolar cyclohexyl rings. Key to attaching the 4'-Ocyclohexyl group was making the 4'-O,6'-O-cyclohexylidene ketal. After peracetylation, the cyclohexylidene ketal ring was opened regioselectively, providing 65% of 8 after final deacetylation. Comparison of the crystal structure of 8, as the cyclohexane solvate, with those of cellulose and its fragments, especially cellotetraose with four glucose rings, revealed extensive effects from the cyclohexyl groups. Three conformationally unique molecules (A, B, and C) are in the triclinic unit cell of 8, along with two solvent cyclohexanes. When viewed down the crystal's a-axis, the array of C, A, and B looks like the letter N, with A inclined so that its cyclohexyl groups can stack with those of the reducing ends of the B and C molecules. The lower left and upper right points of the N are stacks of cyclohexyl rings on the nonreducing ends of B and C, interspersed with solvent cyclohexanes. Whereas cellotetraose has antiparallel (up-down) packing, A and B in 8 are oriented "down" in the unit cell while C is "up". "Down-down-up" (or, alternatively, "up-up-down") packing is rare for carbohydrates. Other unusual details include 06 in all three staggered orientations: one is tg, two are gg, and three are gt, confirmed with CP/MAS 13C NMR. The tg O6 donates a proton to an intramolecular hydrogen bond to O2', opposite to the major schemes in native cellulose I. A similar but novel O6B-H...O2'B hydrogen bond is based on a slightly distorted gg orientation. The hydrogen bonds between parallel molecules are unique, with linkages between O2'A and O2'B, O3'A and O3'B, and O6A and O6B. Other details, such as the bifurcated O3...O5' and ...O6' hydrogen bonds are similar to those of other cellulosic structures. C-H...O hydrogen bonds are extensive along the [110] line of quarter-staggering. The unusual features described here expand the range of structural motifs to be considered for as-yet undetermined cellulose structures.

• Hemoglobin enhances the biological activity of synthetic and natural bacterial (endotoxic) virulence factors: a general principle.

  Authors: Jörg Howe, Walter Richter, Lynn Hawkins, Manfred Rössle, Christian Alexander, Karin Fournier, Jean-Pierre Mach, Thierry Waelli, Reginald M Gorczynski, Artur J Ulmer, Helmut Brade, Alla Zamyatina, Paul Kosma, Ernst Th Rietschel, Klaus Brandenburg

  Medicinal chemistry (Shāriqah (United Arab Emirates)). 12/2008; 4(6):520-5.

  Although hemoglobin (Hb) is mainly present in the cytoplasm of erythrocytes (red blood cells), lower concentrations of pure, cell-free Hb are released permanently into the circulation due to anAlthough hemoglobin (Hb) is mainly present in the cytoplasm of erythrocytes (red blood cells), lower concentrations of pure, cell-free Hb are released permanently into the circulation due to an inherent intravascular hemolytic disruption of erythrocytes. Previously it was shown that the interaction of Hb with bacterial endotoxins (lipopolysaccharides, LPS) results in a significant increase of the biological activity of LPS. There is clear evidence that the enhancement of the biological activity of LPS by Hb is connected with a disaggregation of LPS. From these findings one questions whether the property to enhance the biological activity of endotoxin, in most cases proven by the ability to increase the cytokine (tumor-necrosis-factor-alpha, interleukins) production in human mononuclear cells, is restricted to bacterial endotoxin or is a more general principle in nature. To elucidate this question, we investigated the interaction of various synthetic and natural virulence (pathogenicity) factors with hemoglobin of human or sheep origin. In addition to enterobacterial R-type LPS a synthetic bacterial lipopeptide and synthetic phospholipid-like structures mimicking the lipid A portion of LPS were analysed. Furthermore, we also tested endotoxically inactive LPS and lipid A compounds such as those from Chlamydia trachomatis. We found that the observations made for endotoxically active form of LPS can be generalized for the other synthetic and natural virulence factors: In every case, the cytokine-production induced by them is increased by the addition of Hb. This biological property of Hb is connected with its physical property to convert the aggregate structures of the virulence factors into one with cubic symmetry, accompanied with a considerable reduction of the size and number of the original aggregates.