Vernon Reinhold - Academia.edu (original) (raw)

Papers by Vernon Reinhold

Proceedings of the National Academy of Sciences, 1978

Identification of the principal aflatoxin B1-DNA adduct formed in vivo in rat liver (high-pressur... more Identification of the principal aflatoxin B1-DNA adduct formed in vivo in rat liver (high-pressure liquid chromatography/carcinogen-nucleic acid interaction/field desorption mass spectrometry

Molecular & cellular proteomics : MCP, 2018

The complexity of snake venoms has long been investigated to explore a myriad of biologically act... more The complexity of snake venoms has long been investigated to explore a myriad of biologically active proteins and peptides that are used for immobilizing or killing prey, and are responsible for the pathological effects observed upon envenomation. Glycosylation is the main post-translational modification (PTM) of viperid venoms but currently there is little understanding of how protein glycosylation impacts the variation of venom proteomes. We have previously reported that Bothrops venom glycoproteomes contain a core of components that markedly define their composition and parallel their phylogenetic classification. Here we extend those observations to eight Bothrops species evaluating the N-glycomes by LC-MS as assigned cartoon structures and detailing those structures separately as methylated analogs using ion-trap mass spectrometry (MSn). Following ion disassembly through multiple steps provided sequence and linkage isomeric details that characterized 52 unique compositions in Bo...

Glycobiology, Jan 30, 2014

Glycophorins C and D are highly glycosylated integral sialoglycoproteins of human red blood cell ... more Glycophorins C and D are highly glycosylated integral sialoglycoproteins of human red blood cell membranes carrying the Gerbich blood group antigens. The O- and N-glycosidic chains of the major erythrocyte glycoprotein (Lisowska 2001, Antigenic properties of human glycophorins - an update. Adv Exp Med Biol, 491:155-169; Tomita and Marchesi 1975, Amino-acid sequence and oligosaccharide attachment sites of human erythrocyte glycophorin. Proc Natl Acad Sci USA, 72:2964-2968.) are well characterized but the structure of GPC N-glycans has remained unknown. This problem became important since it was reported that GPC N-glycans play an essential role in the interaction with Plasmodium falciparum EBA-140 merozoite ligand. The elucidation of these structures seems essential for full characterization of the GPC binding site for the EBA-140 ligand. We have employed detailed structural analysis using sequential mass spectrometry to show that many GPC N-glycans contain H2 antigen structures and ...

Blood

Platelets have the shortest shelf-life of all major blood components and are the most difficult t... more Platelets have the shortest shelf-life of all major blood components and are the most difficult to store complicating platelet transfusion practices. Transfused fresh radiolabeled autologous platelets differ significantly in recovery and survival among healthy subjects, however the cause of the inter-individual differences remains unclear. We demonstrated that the loss of sialic acid from the surfaces of cold-stored and transfused platelets promotes their clearance by Ashwell Morell receptors. The loss of platelet surface sialic acid correlates with increases in surface sialidase activity during platelet storage. Here we investigated whether fresh platelets from individual donors exhibit differences in surface sialidase expression and glycan exposure and sialic acid content changes with storage. Methods Platelets were isolated by standard methods from the venous blood of healthy volunteers or from standard platelet concentrates (PCs) and analyzed by flow cytometry for surface β-gala...

The Journal of biological chemistry, Jan 11, 2018

In humans, six α(1,3)-fucosyltransferases (α(1,3)-FTs: FT3/FT4/FT5/FT6/FT7/FT9) reportedly fucosy... more In humans, six α(1,3)-fucosyltransferases (α(1,3)-FTs: FT3/FT4/FT5/FT6/FT7/FT9) reportedly fucosylate terminal lactosaminyl glycans yielding Lewis-X (Le; CD15) and/or sialyl Lewis-X (sLe; CD15s), structures that play key functions in cell migration, development, and immunity. Prior studies analyzing α(1,3)-FT specificities utilized either purified and/or recombinant enzymes to modify synthetic substrates under nonphysiological reaction conditions or molecular biology approaches wherein α(1,3)-FTs were expressed in mammalian cell lines, notably excluding investigations using primary human cells. Accordingly, although significant insights into α(1,3)-FT catalytic properties have been obtained, uncertainty persists regarding their human Le/sLe biosynthetic range across various glycoconjugates. Here, we undertook a comprehensive evaluation of the lactosaminyl product specificities of intracellularly expressed α(1,3)-FTs using a clinically relevant primary human cell type, mesenchymal st...

ABSTRACT Analytical instrumentation and methodology is presented for the determination of endotox... more ABSTRACT Analytical instrumentation and methodology is presented for the determination of endotoxin-related structures at much improved sensitivity and specificity using techniques in chromatography and mass spectrometry. These techniques include a newly synthesized reagent that has provided femtomole sensitivity for oligosaccharide detection; and a preanalysis chemical approach for the determination of oligosaccharide linkage, branching, and sequence. These approaches are compatible with supercritical fluid chromatography-mass spectrometry (SFC-MS) and combine to provide the elements of a global approach to oligosaccharide structure. The utility of this approach has been demonstrated for glycolipids and N-linked glycans. The lipid-A structure from Salmonella minnesota has been demonstrated and the report outlines the currently understood structure Coxiella burnetii lipid-A.

Glycobiology, 2017

Glycosyltransferases, usually residing within the intracellular secretory apparatus, also circula... more Glycosyltransferases, usually residing within the intracellular secretory apparatus, also circulate in the blood. Many of these blood-borne glycosyltransferases are associated with pathological states, including malignancies and inflammatory conditions. Despite the potential for dynamic modifications of glycans on distal cell surfaces and in the extracellular milieu, the glycan-modifying activities present in systemic circulation have not been systematically examined. Here, we describe an evaluation of blood-borne sialyl-, galactosyl- and fucosyltransferase activities that act upon the four common terminal glycan precursor motifs, GlcNAc monomer, Gal(β3)GlcNAc, Gal(β4)GlcNAc and Gal(β3)GalNAc, to produce more complex glycan structures. Data from radioisotope assays and detailed product analysis by sequential tandem mass spectrometry show that blood has the capacity to generate many of the well-recognized and important glycan motifs, including the Lewis, sialyl-Lewis, H- and Sialyl-T...

Discovering the subtleties of sugars : proceedings of the 3rd Beilstein Glyco-Bioinformatics Symposium : June 10th - 14th, 2013, Potsdam, Germany / edited by Martin G. Hicks and Carsten Kettner. International Beilstein Symposium on Glyc..., Jan 22, 2014

Sequential disassembly (MSn) has been applied to fully characterise and document native samples c... more Sequential disassembly (MSn) has been applied to fully characterise and document native samples containing glycan epitopes with their synthetic analogues. Both sample types were prepared by methylation, solvent phase extracted, directly infused and spatially resolved. Product ions of all samples were compiled and contrasted using management tools prepared for the fragment ion library. Each of the epitopes was further disassembled to confirm the multiple structural isomers probable within component substructures of linkage and branching. All native samples tested proved to be matched with their synthetic analogues and reasonably identical on either linear or cylindrical ion traps. Not surprisingly, spectra of mixed epitopes fragment independently, being uninfluenced by similarities. The approach has been coupled with computational tools for data handling and presentation.

Analytical Biochemistry, Apr 30, 1994

Previous studies have defined specific functional relationships within monophosphoryl lipid A (ML... more Previous studies have defined specific functional relationships within monophosphoryl lipid A (MLA) preparations. To extend this understanding to all contributing entities, MLA samples have been structurally characterized using electrospray ionization, collision-induced dissociation (CID), and tandem mass spectrometry (MS/MS). MLA profiles of Salmonella minnesota Re595 have been compared with Shigella flexneri for sample type and component distribution. In excess of 20 individual structures compose each sample, which differ considerably in abundance but little in composition. Component heterogeneity can be directly related to alkane chain length, "lipid X"-type analogs, and variations in esterification to the core 2-amino-2-deoxydisaccharide, GlcNH2 beta(1-6)GlcNH2. The previously defined heptaacyl structure in S. minnesota Re595 was identified at only 15% with the most abundant species a hexaacyl analog. Profiles of S. flexneri MLA show an absence of any heptaacyl analog, with the pentaacyl component the most abundant. To confirm structural relationships, ions from both samples were fragmented by CID and separated by MS/MS. Product ion spectra proved to be identical, showing a series of acyl losses and glycosidic cleavage fragments. Since the position of each acyl group has been previously established in S. minnesota Re595 MLA, ions in the CID spectrum of S. flexneri sample could be structurally assigned. Knowledge of these structural details, their isolation, and biological testing may provide components of unique immune adjuvancy or block-selective deleterious endotoxic processes.

Analytical Biochemistry, Mar 31, 1991

N-Linked glycans have been characterized by supercritical fluid chromatography (SFC) and SFC-MS u... more N-Linked glycans have been characterized by supercritical fluid chromatography (SFC) and SFC-MS using positive- and negative-ion chemical ionization. Four common oligosaccharide derivatives have been prepared and their chromatographic properties assessed on three SFC columns of varying polarity. Carbon dioxide has been used as the SFC mobile phase, with ammonia or CO2 added to the ion source for positive- and negative-ion chemical ionization, respectively. Direct SFC-MS interfacing allows the analytical manipulations of single-ion monitoring, total-ion plots, background subtraction, library searches, and spectral reconstruction algorithms. Positive ammonia chemical ionization yields abundant molecular-weight information, (MH)+, and (MNH4)+ with little or no fragmentation. To capitalize on sensitivity, samples were prepared with the pentafluorobenzyl aminobenzoate reagent, acetylated, and analyzed by SFC-NICI-MS. This modification improves column efficiency and resolution and greatly enhances detecting sensitivity. These "soft" ionization conditions provide abundant molecular-weight-related anions for collision-induced dissociation and subpicogram detection.

Science, Jan 3, 1986

The cyclic (1----2)-beta-D-glucans produced by species of Agrobacterium and Rhizobium resemble th... more The cyclic (1----2)-beta-D-glucans produced by species of Agrobacterium and Rhizobium resemble the membrane-derived oligosaccharides of Escherichia coli in their periplasmic localization, intermediate size, and (1----2)-beta-D-glucan backbones. The regulation of the biosynthesis of cyclic (1----2)-beta-D-glucan by Agrobacterium tumefaciens is now shown to parallel the osmotic regulation of membrane-derived oligosaccharide biosynthesis in Escherichia coli. This result suggests a general role for periplasmic oligosaccharides in the osmotic adaptation of Gram-negative bacteria as ecologically diverse as enteric and soil bacteria.

Glycoconjugate Journal, Aug 1, 1996

We have reported that transglycosylation activity of endo-/3-N-acetylglucosaminidase from Arthrob... more We have reported that transglycosylation activity of endo-/3-N-acetylglucosaminidase from Arthrobacter protophormiae (endo-A) can be enhanced to near completion using GlcNAc as an acceptor in a medium containing 30% acetone (

Proceedings of the National Academy of Sciences, 1978

Identification of the principal aflatoxin B1-DNA adduct formed in vivo in rat liver (high-pressur... more Identification of the principal aflatoxin B1-DNA adduct formed in vivo in rat liver (high-pressure liquid chromatography/carcinogen-nucleic acid interaction/field desorption mass spectrometry

Molecular & cellular proteomics : MCP, 2018

The complexity of snake venoms has long been investigated to explore a myriad of biologically act... more The complexity of snake venoms has long been investigated to explore a myriad of biologically active proteins and peptides that are used for immobilizing or killing prey, and are responsible for the pathological effects observed upon envenomation. Glycosylation is the main post-translational modification (PTM) of viperid venoms but currently there is little understanding of how protein glycosylation impacts the variation of venom proteomes. We have previously reported that Bothrops venom glycoproteomes contain a core of components that markedly define their composition and parallel their phylogenetic classification. Here we extend those observations to eight Bothrops species evaluating the N-glycomes by LC-MS as assigned cartoon structures and detailing those structures separately as methylated analogs using ion-trap mass spectrometry (MSn). Following ion disassembly through multiple steps provided sequence and linkage isomeric details that characterized 52 unique compositions in Bo...

Glycobiology, Jan 30, 2014

Glycophorins C and D are highly glycosylated integral sialoglycoproteins of human red blood cell ... more Glycophorins C and D are highly glycosylated integral sialoglycoproteins of human red blood cell membranes carrying the Gerbich blood group antigens. The O- and N-glycosidic chains of the major erythrocyte glycoprotein (Lisowska 2001, Antigenic properties of human glycophorins - an update. Adv Exp Med Biol, 491:155-169; Tomita and Marchesi 1975, Amino-acid sequence and oligosaccharide attachment sites of human erythrocyte glycophorin. Proc Natl Acad Sci USA, 72:2964-2968.) are well characterized but the structure of GPC N-glycans has remained unknown. This problem became important since it was reported that GPC N-glycans play an essential role in the interaction with Plasmodium falciparum EBA-140 merozoite ligand. The elucidation of these structures seems essential for full characterization of the GPC binding site for the EBA-140 ligand. We have employed detailed structural analysis using sequential mass spectrometry to show that many GPC N-glycans contain H2 antigen structures and ...

Blood

Platelets have the shortest shelf-life of all major blood components and are the most difficult t... more Platelets have the shortest shelf-life of all major blood components and are the most difficult to store complicating platelet transfusion practices. Transfused fresh radiolabeled autologous platelets differ significantly in recovery and survival among healthy subjects, however the cause of the inter-individual differences remains unclear. We demonstrated that the loss of sialic acid from the surfaces of cold-stored and transfused platelets promotes their clearance by Ashwell Morell receptors. The loss of platelet surface sialic acid correlates with increases in surface sialidase activity during platelet storage. Here we investigated whether fresh platelets from individual donors exhibit differences in surface sialidase expression and glycan exposure and sialic acid content changes with storage. Methods Platelets were isolated by standard methods from the venous blood of healthy volunteers or from standard platelet concentrates (PCs) and analyzed by flow cytometry for surface β-gala...

The Journal of biological chemistry, Jan 11, 2018

In humans, six α(1,3)-fucosyltransferases (α(1,3)-FTs: FT3/FT4/FT5/FT6/FT7/FT9) reportedly fucosy... more In humans, six α(1,3)-fucosyltransferases (α(1,3)-FTs: FT3/FT4/FT5/FT6/FT7/FT9) reportedly fucosylate terminal lactosaminyl glycans yielding Lewis-X (Le; CD15) and/or sialyl Lewis-X (sLe; CD15s), structures that play key functions in cell migration, development, and immunity. Prior studies analyzing α(1,3)-FT specificities utilized either purified and/or recombinant enzymes to modify synthetic substrates under nonphysiological reaction conditions or molecular biology approaches wherein α(1,3)-FTs were expressed in mammalian cell lines, notably excluding investigations using primary human cells. Accordingly, although significant insights into α(1,3)-FT catalytic properties have been obtained, uncertainty persists regarding their human Le/sLe biosynthetic range across various glycoconjugates. Here, we undertook a comprehensive evaluation of the lactosaminyl product specificities of intracellularly expressed α(1,3)-FTs using a clinically relevant primary human cell type, mesenchymal st...

ABSTRACT Analytical instrumentation and methodology is presented for the determination of endotox... more ABSTRACT Analytical instrumentation and methodology is presented for the determination of endotoxin-related structures at much improved sensitivity and specificity using techniques in chromatography and mass spectrometry. These techniques include a newly synthesized reagent that has provided femtomole sensitivity for oligosaccharide detection; and a preanalysis chemical approach for the determination of oligosaccharide linkage, branching, and sequence. These approaches are compatible with supercritical fluid chromatography-mass spectrometry (SFC-MS) and combine to provide the elements of a global approach to oligosaccharide structure. The utility of this approach has been demonstrated for glycolipids and N-linked glycans. The lipid-A structure from Salmonella minnesota has been demonstrated and the report outlines the currently understood structure Coxiella burnetii lipid-A.

Glycobiology, 2017

Glycosyltransferases, usually residing within the intracellular secretory apparatus, also circula... more Glycosyltransferases, usually residing within the intracellular secretory apparatus, also circulate in the blood. Many of these blood-borne glycosyltransferases are associated with pathological states, including malignancies and inflammatory conditions. Despite the potential for dynamic modifications of glycans on distal cell surfaces and in the extracellular milieu, the glycan-modifying activities present in systemic circulation have not been systematically examined. Here, we describe an evaluation of blood-borne sialyl-, galactosyl- and fucosyltransferase activities that act upon the four common terminal glycan precursor motifs, GlcNAc monomer, Gal(β3)GlcNAc, Gal(β4)GlcNAc and Gal(β3)GalNAc, to produce more complex glycan structures. Data from radioisotope assays and detailed product analysis by sequential tandem mass spectrometry show that blood has the capacity to generate many of the well-recognized and important glycan motifs, including the Lewis, sialyl-Lewis, H- and Sialyl-T...

Discovering the subtleties of sugars : proceedings of the 3rd Beilstein Glyco-Bioinformatics Symposium : June 10th - 14th, 2013, Potsdam, Germany / edited by Martin G. Hicks and Carsten Kettner. International Beilstein Symposium on Glyc..., Jan 22, 2014

Sequential disassembly (MSn) has been applied to fully characterise and document native samples c... more Sequential disassembly (MSn) has been applied to fully characterise and document native samples containing glycan epitopes with their synthetic analogues. Both sample types were prepared by methylation, solvent phase extracted, directly infused and spatially resolved. Product ions of all samples were compiled and contrasted using management tools prepared for the fragment ion library. Each of the epitopes was further disassembled to confirm the multiple structural isomers probable within component substructures of linkage and branching. All native samples tested proved to be matched with their synthetic analogues and reasonably identical on either linear or cylindrical ion traps. Not surprisingly, spectra of mixed epitopes fragment independently, being uninfluenced by similarities. The approach has been coupled with computational tools for data handling and presentation.

Analytical Biochemistry, Apr 30, 1994

Previous studies have defined specific functional relationships within monophosphoryl lipid A (ML... more Previous studies have defined specific functional relationships within monophosphoryl lipid A (MLA) preparations. To extend this understanding to all contributing entities, MLA samples have been structurally characterized using electrospray ionization, collision-induced dissociation (CID), and tandem mass spectrometry (MS/MS). MLA profiles of Salmonella minnesota Re595 have been compared with Shigella flexneri for sample type and component distribution. In excess of 20 individual structures compose each sample, which differ considerably in abundance but little in composition. Component heterogeneity can be directly related to alkane chain length, "lipid X"-type analogs, and variations in esterification to the core 2-amino-2-deoxydisaccharide, GlcNH2 beta(1-6)GlcNH2. The previously defined heptaacyl structure in S. minnesota Re595 was identified at only 15% with the most abundant species a hexaacyl analog. Profiles of S. flexneri MLA show an absence of any heptaacyl analog, with the pentaacyl component the most abundant. To confirm structural relationships, ions from both samples were fragmented by CID and separated by MS/MS. Product ion spectra proved to be identical, showing a series of acyl losses and glycosidic cleavage fragments. Since the position of each acyl group has been previously established in S. minnesota Re595 MLA, ions in the CID spectrum of S. flexneri sample could be structurally assigned. Knowledge of these structural details, their isolation, and biological testing may provide components of unique immune adjuvancy or block-selective deleterious endotoxic processes.

Analytical Biochemistry, Mar 31, 1991

N-Linked glycans have been characterized by supercritical fluid chromatography (SFC) and SFC-MS u... more N-Linked glycans have been characterized by supercritical fluid chromatography (SFC) and SFC-MS using positive- and negative-ion chemical ionization. Four common oligosaccharide derivatives have been prepared and their chromatographic properties assessed on three SFC columns of varying polarity. Carbon dioxide has been used as the SFC mobile phase, with ammonia or CO2 added to the ion source for positive- and negative-ion chemical ionization, respectively. Direct SFC-MS interfacing allows the analytical manipulations of single-ion monitoring, total-ion plots, background subtraction, library searches, and spectral reconstruction algorithms. Positive ammonia chemical ionization yields abundant molecular-weight information, (MH)+, and (MNH4)+ with little or no fragmentation. To capitalize on sensitivity, samples were prepared with the pentafluorobenzyl aminobenzoate reagent, acetylated, and analyzed by SFC-NICI-MS. This modification improves column efficiency and resolution and greatly enhances detecting sensitivity. These "soft" ionization conditions provide abundant molecular-weight-related anions for collision-induced dissociation and subpicogram detection.

Science, Jan 3, 1986

The cyclic (1----2)-beta-D-glucans produced by species of Agrobacterium and Rhizobium resemble th... more The cyclic (1----2)-beta-D-glucans produced by species of Agrobacterium and Rhizobium resemble the membrane-derived oligosaccharides of Escherichia coli in their periplasmic localization, intermediate size, and (1----2)-beta-D-glucan backbones. The regulation of the biosynthesis of cyclic (1----2)-beta-D-glucan by Agrobacterium tumefaciens is now shown to parallel the osmotic regulation of membrane-derived oligosaccharide biosynthesis in Escherichia coli. This result suggests a general role for periplasmic oligosaccharides in the osmotic adaptation of Gram-negative bacteria as ecologically diverse as enteric and soil bacteria.

Glycoconjugate Journal, Aug 1, 1996

We have reported that transglycosylation activity of endo-/3-N-acetylglucosaminidase from Arthrob... more We have reported that transglycosylation activity of endo-/3-N-acetylglucosaminidase from Arthrobacter protophormiae (endo-A) can be enhanced to near completion using GlcNAc as an acceptor in a medium containing 30% acetone (