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D Oxley

Publications and source records attributed to D Oxley.

At least 19 recordsLinked to original sources
PubMed2001

NaAGP4 is an arabinogalactan protein whose expression is suppressed by wounding and fungal infection in Nicotiana alata.

Arabinogalactan proteins (AGPs) are proteoglycans secreted by plant cells that have been implicated in plant growth and development. Most AGPs cloned to date possess highly labile glycosylphosphatidylinositol (GPI) lipid anchors. These anchors transiently attach AGPs to the plasma membrane before they are released into the cell wall following GPI anchor hydrolysis. We have isolated and partially sequenced the protein core of an AGP purified from styles of Nicotiana alata. The protein sequence data were utilised to clone the AGP's gene, NaAGP4. This AGP shares about 78% sequence identity with the tomato AGP LeAGP-1. RNA gel blot analyses of different plant organs indicate that NaAGP4 is expressed in the same tissues and at similar levels as LeAGP-1. Furthermore, NaAGP4 like LeAGP-1 is rapidly suppressed by tissue wounding and by pathogen infection. We believe NaAGP4 and LeAGP-1 are the first described examples of orthologous AGPs from different plant species. In contrast, another AGP from N. alata, NaAGP1, is comparatively unaffected by wounding and pathogen infection, although this AGP is expressed in similar tissues and at similar levels as NaAGP4.

Amino Acid Sequence↗
PubMed1999

Structure of the glycosylphosphatidylinositol anchor of an arabinogalactan protein from Pyrus communis suspension-cultured cells.

Arabinogalactan proteins (AGPs) are proteoglycans of higher plants, which are implicated in growth and development. We recently have shown that two AGPs, NaAGP1 (from Nicotiana alata styles) and PcAGP1 (from Pyrus communis cell suspension culture), are modified by the addition of a glycosylphosphatidylinositol (GPI) anchor. However, paradoxically, both AGPs were buffer soluble rather than membrane associated. We now show that pear suspension cultured cells also contain membrane-bound GPI-anchored AGPs. This GPI anchor has the minimal core oligosaccharide structure, D-Manalpha(1-2)-D-Manalpha(1-6)-D-Manalpha(1-4)-D-GlcN -inositol, which is consistent with those found in animals, protozoa, and yeast, but with a partial beta(1-4)-galactosyl substitution of the 6-linked Man residue, and has a phosphoceramide lipid composed primarily of phytosphingosine and tetracosanoic acid. The secreted form of PcAGP1 contains a truncated GPI lacking the phosphoceramide moiety, suggesting that it is released from the membrane by the action of a phospholipase D. The implications of these findings are discussed in relation to the potential mechanisms by which GPI-anchored AGPs may be involved in signal transduction pathways.

Cells, Cultured↗
PubMed1998

Arabinogalactan-proteins from Nicotiana alata and Pyrus communis contain glycosylphosphatidylinositol membrane anchors.

Arabinogalactan-proteins (AGPs) are a class of proteoglycans found in cell secretions and plasma membranes of plants. Attention is currently focused on their structure and their potential role in growth and development. We present evidence that two members of a major class of AGPs, the classical AGPs, AGPNa1 from styles of Nicotiana alata and AGPPc1 from cell suspension cultures of Pyrus communis, undergo C-terminal processing involving glycosylphosphatidylinositol membrane anchors. The evidence is that (i) the transmembrane helix at the C terminus predicted from the cDNA encoding these proteins is not present-the C-terminal amino acid is Asn87 and Ser97 for AGPNa1 and AGPPc1, respectively; (ii) both AGP protein backbones are substituted with ethanolamine at the C-terminal amino acid; and (iii) inositol, glucosamine, and mannose are present in the native AGPs. An examination of the deduced amino acid sequences of other classical AGP protein backbones shows that glycosylphosphatidylinositol-anchors may be a common feature of this class of AGPs.

Amino Acid Sequence↗
PubMed1998

Structure and distribution of N-glycans on the S7-allele stylar self-incompatibility ribonuclease of Nicotiana alata.

S-RNases are the stylar products of the self-incompatibility (S)-locus in solanaceous plants (including Nicotiana alata), and as such, are involved in the prevention of self-pollination. All cDNA sequences of S-RNase products of functional S-alleles contain potential N-glycosylation sites, with one site being conserved in all cases, suggesting that N-glycosylation is important in self-incompatibility. In this study, we report on the structure and localization of the N-glycans on the S7-allele RNase of N. alata. A total of nine N-glycans, belonging to the high-mannose- and xylosylated hybrid-classes, were identified and characterized by a combination of electrospray-ionization mass-spectrometry (ESI-MS), 1H-NMR spectroscopy, and methylation analyses. The glycosylation pattern of individual glycosylation sites was determined by ESI-MS of the glycans released from isolated chymotryptic glycopeptides. All three N-glycosylation sites showed microheterogeneity and each had a unique complement of N-glycans. The N-glycosylation pattern of the S7-RNase is significantly different to those of the S1- and S2-RNases.

Alleles↗
PubMed1998

Structural analysis and molecular model of a self-incompatibility RNase from wild tomato.

Self-incompatibility RNases (S-RNases) are an allelic series of style glycoproteins associated with rejection of self-pollen in solanaceous plants. The nucleotide sequences of S-RNase alleles from several genera have been determined, but the structure of the gene products has only been described for those from Nicotiana alata. We report on the N-glycan structures and the disulfide bonding of the S3-RNase from wild tomato (Lycopersicon peruvianum) and use this and other information to construct a model of this molecule. The S3-RNase has a single N-glycosylation site (Asn-28) to which one of three N-glycans is attached. S3-RNase has seven Cys residues; six are involved in disulfide linkages (Cys-16-Cys-21, Cys-46-Cys-91, and Cys-166-Cys-177), and one has a free thiol group (Cys-150). The disulfide-bonding pattern is consistent with that observed in RNase Rh, a related RNase for which radiographic-crystallographic information is available. A molecular model of the S3-RNase shows that four of the most variable regions of the S-RNases are clustered on one surface of the molecule. This is discussed in the context of recent experiments that set out to determine the regions of the S-RNase important for recognition during the self-incompatibility response.

Amino Acid Sequence↗
PubMed1997

Identification of active-site histidine residues of a self-incompatibility ribonuclease from a wild tomato.

The style component of the self-incompatibility (S) locus of the wild tomato Lycopersicon peruvianum (L.) Mill. is an allelic series of glycoproteins with ribonuclease activity (S-RNases). Treatment of the S3-RNase from L. peruvianum with iodoacetate at pH 6.1 led to a loss of RNase activity. In the presence of a competitive inhibitor, guanosine 3'-monophosphate (3'-GMP), the rate of RNase inactivation by iodoacetate was reduced significantly. Analysis of the tryptic digestion products of the iodoacetate-modified S-RNase by reversed-phase high-performance liquid chromatography and electrospray-ionization mass spectrometry showed that histidine-32 was preferentially modified in the absence of 3'-GMP. Histidine-88 was also modified, but this occurred both in the presence and absence of 3'-GMP, suggesting that this residue is accessible when 3'-GMP is in the active site. Cysteine-150 was modified by iodoacetate in the absence of 3'-GMP and, to a lesser extent, in its presence. The results are discussed with respect to the related fungal RNase T2 family and the mechanism of S-RNase action.

Amino Acid Sequence↗
PubMed1996

Disulphide bonding in a stylar self-incompatibility ribonuclease of Nicotiana alata.

Many flowering plants have developed a self-incompatibility mechanism, which is controlled by a single polyallelic locus (the S-locus), to prevent inbreeding. The products of the S-locus in the styles of solanaceous plants are an allelic series of glycoproteins with RNase activity [McClure, B. A., Haring, V., Ebert, P. R., Anderson, M. A., Simpson, R. J., Sakiyama, F. & Clarke, A. E. (1989) Nature 342, 955-957]. These S-RNases show some amino-acid-sequence similarity with two fungal RNases (T2 and Rh), including the presence of two active-site His residues, which suggests a common three-dimensional structure. Disulphide bonding is important in the maintenance of the three-dimensional structure of the fungal RNases [Kurihara, H., Mitsui, Y., Ohgi, K., Irie, M., Mizuno, H. & Nakamura, T. (1992) FEBS Lett. 306, 189-192] and the S-RNases [Tsai, D. S., Lee, H.-S., Post, L. C., Kreiling, K. M. & Kao, T.-H. (1992) Sex. Plant Reprod. 5, 256-263]. We have used the S2-allele RNase of Nicotiana alata, which has nine Cys residues, to establish the pattern of disulphide bonding. The disulphide bonds Cys16-Cys21, Cys45-Cys94, Cys153-Cys182 and Cys165-Cys176 are consistent with the S2-RNase having a similar three-dimensional structure to RNase Rh. A free Cys residue (Cys95) adjacent to Cys45-Cys94 promotes a rapid specific disulphide migration when the protein is exposed to denaturing conditions.

Amino Acid Sequence↗
PubMed1996

Structure of N-glycans on the S3- and S6-allele stylar self-incompatibility ribonucleases of Nicotiana alata.

Self-incompatibility is a mechanism developed by many plants to prevent inbreeding. The products of the self-incompatibility (S)-locus in the styles of solanaceous plants are a series of glycoproteins with ribonuclease activity. In this study, we report on the N-glycans from the stylar self-incompatibility S3- and S6-ribonucleases of Nicotiana alata, which were enzymically released and fractionated by high-pH anion-exchange HPLC. A total of 14 N-glycans were identified and characterized by a combination of electrospray-ionization mass-spectrometry, 1H-NMR spectros-copy, chemical degradation, and methylation analyses. This patterns of N-glycosylation is much more complex than that previously found on the N.alata S1- and S2-RNases, each of which contained only four N-glycans.

Alleles↗
PubMed1995

Structure of the N-linked oligosaccharides from tridacnin, a lectin found in the haemolymph of the giant clam Hippopus hippopus.

Tridacnin, a glycoprotein lectin, was isolated from the symbiotic marine clam Hippopus hippopus and the structure of its major N-glycan chains determined. Tridacnin contains only N-linked glycans which were quantitatively cleaved by peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F. Following purification by anion-exchange HPLC, the structures of the oligosaccharides were established using a combination of electrospray ionisation mass spectrometry, 1H-NMR spectroscopy and linkage analysis. The N-glycans are primarily of the oligomannose type but, in addition, some contain a novel 6-O-Me group on the terminal mannose residue of the chain. The N-glycan chains had the following structures. [formula: see text]

Animals↗
PubMed1995

Microheterogeneity of N-glycosylation on a stylar self-incompatibility glycoprotein of Nicotiana alata.

Gametophytic self-incompatibility, a mechanism that prevents inbreeding in some families of flowering plants, is mediated by the products of a single genetic locus, the S-locus. The products of the S-gene in the female sexual tissues of Nicotiana alata are an allelic series of glycoproteins with RNase activity. In this study, we report on the microheterogeneity of N-linked glycosylation at the four potential N-glycosylation sites of the S2-glycoprotein. The S-glycoproteins from N.alata contain from one to five potential N-glycosylation sites based on the consensus sequence Asn-Xaa-Ser/Thr. The S2-glycoprotein contains four potential N-glycosylation sites at Asn27, Asn37, Asn38 and Asn 150, designated sites I, II, IV and V, respectively. Site III is absent from the S2-glycoprotein. Analysis of glycopeptides generated from the S2-glycoprotein by trypsin and chymotrypsin digestions revealed the types of glycans and the degree of microheterogeneity present at each site. Sites I (Asn27) and IV (Asn138) display microheterogeneity, site II (Asn37) contains only a single type of N-glycan, and site V (Asn150) is not glycosylated. The microheterogeneity observed at site I on the S2-glycoprotein is the same as that observed at the only site, site I, on the S1-glycoprotein (Woodward et al., Glycobiology, 2, 241-250, 1992). Since the N-glycosylation consensus sequence at site I is conserved in all S-glycoproteins from other species of self-incompatible solanaceous plants, glycosylation at this site may be important to their function. No other post-translational modifications (e.g. O-glycosylation, phosphorylation) were detected on the S2-glycoprotein.

Alleles↗
PubMed1993

Structural and serological characterisation of an O-specific polysaccharide from Serratia plymuthica.

The surface polysaccharides of a strain of Serratia plymuthica were characterised and shown to consist of a linear, acidic galactoglucomannan as well as a major and a minor neutral galactan. Immunoblotting results demonstrated cross-reactions between this strain and others with similar galactans (S. marcescens O16 and O20, Klebsiella O1, and Pasteurella haemolytica T4 and T10).

Antigens, Bacterial↗
PubMed1992

Structure of a neutral glycan from the lipopolysaccharides of reference strains for Serratia marcescens serogroups O2 and O3.

Serogroups O2 and O3 of Serratia marcescens are differentiated by acidic glycans present in the aqueous phase when lipopolysaccharides are extracted from the reference strains by the aqueous-phenol method. The phenolic phases of these extracts from both strains also contain lipopolysaccharides, from which the same neutral glycan is released on milk acid hydrolysis. The neutral glycan has the disaccharide repeating-unit shown, and accounts for the cross-reactions between the two serogroups and also with serogroup O21: --> 4)-alpha-D-GlcpNAc-(1-->4)-beta-D-ManpNAc-(1--.

Carbohydrate Sequence↗
PubMed1992

Structure of an acidic glycan from the reference strain for Serratia marcescens serogroup O22.

In addition to a neutral glycan, lipopolysaccharide extracts from the reference strain for Serratia marcescens serogroup O22 contain an acidic polymer which probably defines the serogroup and is of microcapsular origin. The polymer is doubly branched with a heptasaccharide repeating unit and a galactan backbone. By means of spectroscopic and degradative studies, the structure of the repeating unit was established as that shown. [formula: see text]

Carbohydrate Conformation↗
PubMed1992

An animal model of fibrinolytic bleeding based on the rebleed phenomenon: application to a study of vulnerability of hemostatic plugs of different age.

The primary bleeding time is prolonged when tested during the infusion of both plasminogen activators and anticoagulants, and such sites frequently exhibit rebleeding after initial hemostatic control. This study describes an animal (rabbit) model which distinguishes fibrinolytic from anticoagulant hemorrhage and further applies the model to the study of hemostatic plugs of increasing age. In this model, rebleeding occurred from hemostatically-stable ear puncture sites induced prior to infusion of streptokinase (SK) or recombinant tissue-plasminogen activator (rt-PA), but not of heparin or hirudin. This distinction was apparent even for lesions induced only 15 minutes prior to the infusion and fibrinolytic bleeding was observed in such lesions induced up to 24 hours earlier. Post-infusion sites bled more quickly than did pre-infusion sites, and there was a gradual decrease in susceptibility of such prior trauma sites for rebleeding, evidenced not only by a lower proportion of sites that rebled, but also by a longer lag time after starting SK or rt-PA before such rebleeding occurred. At the dosages tested, SK showed a trend (not statistically significant) toward more sites that rebled, while rt-PA showed a trend towards a longer duration of rebleeding. Thus, this animal model of rebleeding appears to be unique for fibrinolytic agents and allows for more detailed study of the physiological mechanisms of such bleeding and for a multifaceted comparison of the bleeding potential of plasminogen activators.

Animals↗
PubMed1991

Structure of an acidic glycan present in the lipopolysaccharide extract from the reference strain for Serratia marcescens serogroup O18.

The lipopolysaccharide extract from the cell wall of the reference strain for Serratia marcescens serogroup O18 contained, in addition to a neutral glycan characterised previously, an acidic glycan. Acidity was contributed both by D-glucuronic acid and by 4-O-[(R)-1-carboxyethyl]-D-glucose (4-O-Lac-D-Glc). By using n.m.r. spectroscopy, methylation analysis, and chemical degradations, the repeating unit of the acidic glycan was identified as a branched hexasaccharide having the structure shown; an O-acetyl group also present was not located. The glycan is believed to define the O18 serogroup, but is probably not an integral component of the lipopolysaccharide. [formula: see text].

Carbohydrate Conformation↗
PubMed1991

Structure of the 021 antigen from Serratia marcescens.

Lipopolysaccharide was isolated from both phases of an aqueous-phenol extraction of defatted cell walls from the reference strain for Serratia marcescens serogroup 021. The product from the aqueous phase was of the R type, lacking a polymeric side-chain. The polymeric fraction of the lipopolysaccharide from the phenolic phase (the 021 antigen) had a disaccharide repeating-unit with the following structure: ----4)-alpha-D-Glcp-(1----4)-beta-D-ManpNAc-(1----.

Antigens, Bacterial↗