Structural Investigations on a Cell-Wall Lipopolysaccharide from Neisseria sicca

1971 ◽  
Vol 49 (2) ◽  
pp. 243-250 ◽  
Author(s):  
G. A. Adams
Keyword(s):  
Lipid A ◽  
Periodate Oxidation ◽  
Amino Groups ◽  
Structural Investigations ◽  
Acid Protein ◽  
A Cell ◽  
Backbone Structure ◽  
Neisseria Sicca ◽  
Fourfold Excess ◽  
Phosphate Groups

Lipopolysaccharide (LPS) prepared from Neisseria sicca in 1.5% yield contained D-glucose, D-glucosamine, D-galactosamine, 3-deoxyoctulosonic acid, protein, lipid A, and phosphate. The molecule was judged to be homogeneous as tested by free boundary electrophoresis. D-Galactosamine was associated exclusively with the polysaccharide portion of the molecule and was in fourfold excess of D-glucosamine. The latter hexosamine was primarily a constituent of the lipid A moiety in which it formed the backbone structure linked glycosidically 1 → 4. To this structure, fatty acids, principally β-hydroxymyristic acid and β-hydroxylauric acid, were linked along with phosphate groups. The D-glucosamine units in the polysaccharide portion of the LPS molecule were also attached by 1 → 4 glycosidic linkages. D-Galactosamine units did not survive the methylation procedures due presumably to the lack of acyl protecting groups on its amino groups. Methylation results showed that approximately one-third of the D-glucose units were nonreducing end groups, approximately one-third were linked α1 → 2, a small proportion was linked 1 → 4, and the remainder was branched through C-3, C-4, and C-6. Periodate oxidation results were in agreement with the structure proposed on the basis of the methylation data. The LPS of N. sicca was considerably simpler than that of N. perflava and lacked heptose, rhamnose, and ethanolamine components.

scholarly journals A murine fer testis-specific transcript (ferT) encodes a truncated Fer protein.

1990 ◽  
Vol 10 (1) ◽  
pp. 146-153 ◽  
Author(s):  
K Fischman ◽  
J C Edman ◽  
G M Shackleford ◽  
J A Turner ◽  
W J Rutter ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Translation System ◽  
Appearance Time ◽  
Acid Protein ◽  
Mouse Tissues ◽  
Free Translation ◽  
Alternatively Spliced ◽  
A Cell ◽  
Testis Cdna ◽  
Carboxy Terminal

A cDNA for a potential tyrosine kinase-encoding mRNA was isolated from a mouse testis cDNA library. In a survey of eight mouse tissues, a transcript of 2.4 kilobases restricted to testis tissue was found. The mRNA encodes a 453-amino-acid protein of 51,383 daltons, the smallest tyrosine kinase protein ever described. RNA synthesized from the cDNA template directs the synthesis of a 51,000-Mr protein in a cell-free translation system. The carboxy-terminal 409 amino acids are 98 and 90% identical to the carboxy halves of the rat and human Fer proteins, respectively. This suggests that the cDNA represents an alternatively spliced testis-specific fer mRNA and is therefore termed by us ferT. On the basis of the appearance time of the fer mRNA in the testis of maturing neonatal mice, we speculate on the role played by this protein in the development of this organ.

1-Deoxyglycitolation of protein amino groups and their regeneration by periodate oxidation*

2009 ◽  
Vol 26 (1) ◽  
pp. 55-62 ◽  
Author(s):  
W.S. DOMINIC WONG ◽  
MAGNUS M. KRISTJANSSON ◽  
DAVID T. OSUGA ◽  
ROBERT E. FEENEY
Keyword(s):  
Periodate Oxidation ◽  
Amino Groups

cDNA cloning, heterologous expression and characterization of a cell wall invertase from copper tolerant population of Elsholtzia haichowensis

Biologia ◽  
2015 ◽  
Vol 70 (8) ◽  
Author(s):  
Chen Liu ◽  
Zhongrui Xu ◽  
Shenwen Cai ◽  
Luan Zhang ◽  
Zhiting Xiong
Keyword(s):  
Cell Wall ◽  
Evolutionary Relationship ◽  
Methylotrophic Yeast ◽  
Cell Wall Invertase ◽  
Reading Frame ◽  
Methylotrophic Yeast Pichia Pastoris ◽  
Acid Protein ◽  
Elsholtzia Haichowensis ◽  
A Cell ◽  
Theoretical Molecular Mass

AbstractThe main objective of the present study was to clone, heterologously express and characterize a novel cell wall invertase (FCWI) from a Cu tolerant population of Elsholtzia haichowensis. The full-length FCWI cDNA contained an open reading frame (ORF) of 1671 bp which encoded a 556-amino-acid protein. The theoretical molecular mass and pI of the deduced protein were 62.5 kDa and 9.29, respectively. Phylogenetic analysis showed that FCWI had a closer evolutionary relationship to cell wall invertase of dicot. FCWI was expressed in methylotrophic yeast Pichia pastoris and purified to near homogeneity. Recombinant FCWI enzyme had pH optima of 4.0 and temperature optima of 50◦C. Activity analyses in the presence of various metal cations indicated that FCWI was completely inhibited by Hg

scholarly journals Endotoxin protein: a B-cell mitogen and polyclonal activator of C3H/HeJ lymphocytes.

1976 ◽  
Vol 144 (3) ◽  
pp. 821-827 ◽  
Author(s):  
B M Sultzer ◽  
G W Goodman
Keyword(s):  
B Cell ◽  
Lipid A ◽  
Protein A ◽  
Cell Wall Protein ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Lipopolysaccharide Endotoxin ◽  
Polyclonal Activator ◽  
A Cell ◽  
Cell Mitogen

A cell wall protein that is ordinarily complexed to the lipopolysaccharide endotoxin in gram-negative bacteria has been separated by the use of aqueous phenol. The protein is active as a B-cell mitogen and polyclonal activator of murine lymphocytes including the C3H/HeJ strain which is a nonresponder to lipoplysaccharide or lipid A.

Structure of the lipopolysaccharide O-chain of Yersinia enterocolitica serotype O:5,27

1987 ◽  
Vol 65 (1) ◽  
pp. 1-7 ◽  
Author(s):  
Malcolm B. Perry ◽  
Leann L. MacLean
Keyword(s):  
Nuclear Magnetic Resonance ◽  
Yersinia Enterocolitica ◽  
Lipid A ◽  
Optical Rotation ◽  
Periodate Oxidation ◽  
Partial Hydrolysis ◽  
Specific Optical Rotation ◽  
Core Oligosaccharide ◽  
Magnetic Resonance Studies ◽  
Serotype O

The cellular lipopolysaccharide produced by Yersinia enterocolitica serotype O:5,27 was of the S-type and composed of an antigenic O-chain polysaccharide linked through a core oligosaccharide region, which in turn was linked through 3-deoxy-D-manno-octulonosyl units to a lipid A moiety. The O-chain polysaccharide was composed of equal molar amounts of L-rhamnose and D-xylulose. By partial hydrolysis, periodate oxidation, methylation, specific optical rotation, and 13C and 1H nuclear magnetic resonance studies, the structure of the O-chain was established as being a linear backbone of alternating 1,3-linked α-L-rhamnopyranosyl and β-L-rhamnopyranosyl units, to which 2,2-linked β-D-threo-pent-2-ulofuranoside (D-xylulofuranoside) units were present on every L-rhamnopyranosyl residue, as shown below.[Formula: see text]

scholarly journals The configuration of 2,6-diamino-3-hydroxypimelic acid in microbial cell walls

1969 ◽  
Vol 115 (4) ◽  
pp. 797-805 ◽  
Author(s):  
H R Perkins
Keyword(s):  
Cell Wall ◽  
Hydroxy Group ◽  
Cell Walls ◽  
Optical Rotation ◽  
Periodate Oxidation ◽  
Ring Closure ◽  
Diaminopimelic Acid ◽  
Amino Group ◽  
Amino Groups ◽  
Peptide Linkage

β-Hydroxydiaminopimelic acid, together with some diaminopimelic acid, occurs in the cell-wall mucopeptide of certain Actinomycetales. These components were converted into their di-DNP derivatives and separated by chromatography. Hence the relative proportions present in the cell walls of a number of species were measured. The problem of acid-induced inversion of configuration was studied. Of the diaminohydroxypimelic acids isomer B (see Scheme 2; amino groups meso, hydroxy group threo to its neighbouring amino group) always predominated but a small proportion of isomer D (amino groups l, hydroxy group erythro) also occurred. The configuration of the diaminohydroxypimelic acids was determined by periodate oxidation to glutamic γ-semialdehyde, which underwent spontaneous ring-closure. Reduction with sodium borohydride produced optically active proline, the configuration of which was determined by direct measurement of the optical rotation of DNP-proline. Un-cross-linked diaminohydroxypimelic acid in the cell wall was oxidized with periodate in the presence of ammonia. Since the remaining amino group was bound in peptide linkage, ring-closure was prevented and borohydride reduction of the aldehyde–ammonia presumed to be present resulted in the formation of ornithine. The quantity of ornithine was used as a measure of the degree of cross-linking.

scholarly journals Endotoxic lipid A induces intracellular Ca2+ increase in human platelets

1991 ◽  
Vol 278 (1) ◽  
pp. 75-80 ◽  
Author(s):  
M Romano ◽  
M Molino ◽  
C Cerletti
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Lipid A ◽  
Human Platelets ◽  
Free System ◽  
Fura 2 ◽  
Kinase C ◽  
Activated Platelets ◽  
A Cell ◽  
Protein Kinase C Activation

The activation of protein kinase C by endotoxic lipid A was observed with both intact platelets and in a cell-free system [Romano & Hawiger (1990) J. Biol. Chem. 265, 1765-1770]. We have now studied the action of lipid A on intracellular Ca2+ concentration ([Ca2+]i). Lipid A induced a concentration-dependent rise in [Ca2+]i in human platelets loaded with fura-2, which reached a maximum at 37.1 +/- 3.8 s (tmax). Maximum [Ca2+]i levels, observed at 30 microM lipid A, were 432 +/- 60 nM. EGTA (2 mM) or NiCl2 (1 mM) each decreased the lipid A-dependent elevation of [Ca2+]i by 50-60% without significant modification of tmax, but shortening the time for 50% recovery (t50) from greater than 400 s to 113.1 +/- 29.1 s and 54 +/- 2.1 s, respectively. Quenching of the fura-2 signal was also observed in lipid A-stimulated platelets resuspended with MnCl2 (1 mM), suggesting that both mobilization and external influx of Ca2+ occur. Intracellular Ca2+ mobilization depended on release from Ins(1,4,5)P3-sensitive stores, since Ins(1,4,5)P3 accumulation was detected in lipid A-activated platelets. Staurosporine, an inhibitor of protein kinase C, blocked the [Ca2+]i rise generated by lipid A in platelets [concn. giving 50% inhibition (IC50) = 0.1 microM], prolonging the tmax. to 54.7 +/- 5.1 s, but decreasing the t50 to 157.5 +/- 31.8 s. Staurosporine also suppressed InsP3 accumulation (IC50 = 0.15 microM). These results suggest that platelet activation by lipid A involves an interaction between [Ca2+]i elevation and protein kinase C activation.

scholarly journals Developmental and spatial patterns of expression of the mouse homeobox gene, Hox 2.1

Development ◽  
1987 ◽  
Vol 99 (4) ◽  
pp. 603-617 ◽  
Author(s):  
R. Krumlauf ◽  
P.W. Holland ◽  
J.H. McVey ◽  
B.L. Hogan
Keyword(s):  
Gene Expression ◽  
Amino Acids ◽  
Spinal Cord ◽  
Homeobox Gene ◽  
Chromosome 11 ◽  
Cell Type Specificity ◽  
Acid Protein ◽  
Visceral Yolk Sac ◽  
Epithelial Cell Layer ◽  
A Cell

The Hox 2.1 gene forms part of a cluster of homeobox-containing genes on mouse chromosome 11. Analysis of Hox 2.1 cDNAs isolated from an 8 1/2-day p.c. mouse embryo library predicts that the gene encodes a 269 amino acid protein (Mr, 29,432). This deduced protein contains a homeobox 15 amino acids from the carboxy terminus and is very rich in serine and proline. A second partially conserved region present in several other genes containing homeoboxes, the hexapeptide Ile-Phe-Pro-Trp-Met-Arg, is located 12 amino acids upstream of the homeodomain and is encoded by a separate exon. Analysis of Hox 2.1 gene expression reveals a complex and tissue-specific series of RNA transcripts in a broad range of fetal tissues (lung, spinal cord, kidney, gut, spleen, liver and visceral yolk sac). Comparison of the temporal patterns of gene expression during development and in the adult suggests that Hox 2.1 is regulated independently in different tissues. Evidence is also presented that transcripts from other loci have extensive homology to the Hox 2.1 gene in sequences outside of the homeobox. In situ hybridization shows that Hox 2.1 transcripts are regionally localized in the spinal cord in an apparent anterior-posterior gradient extending from the hind brain. The distribution of RNA also displays a cell-type specificity in the lung, where mesodermal cells surrounding the branching epithelial cell layer accumulate high levels of Hox 2.1 transcripts.

scholarly journals Transdermal Vaccination with the Matrix-2 Protein Virus-like Particle (M2e VLP) Induces Immunity in Mice against Influenza A Virus

Vaccines ◽  
2021 ◽  
Vol 9 (11) ◽  
pp. 1324
Author(s):  
Kimberly Braz Gomes ◽  
Sucheta D’Sa ◽  
Grace Lovia Allotey-Babington ◽  
Sang-Moo Kang ◽  
Martin J. D’Souza
Keyword(s):  
T Cells ◽  
Lymph Node ◽  
Influenza A ◽  
Lipid A ◽  
Adaptive Immune ◽  
Monophosphoryl Lipid A ◽  
Virus Like Particle ◽  
The Matrix ◽  
A Cell ◽  
Potential Vaccine

In this study, our goal was to utilize the extracellular domain matrix-2 protein virus-like particle (M2e VLP) that has been found to be highly conserved amongst all strains of influenza and could serve as a potential vaccine candidate against influenza. Previous studies have demonstrated that the VLP of the M2e showed increased activation of innate and adaptive immune responses. Therefore, to further explore its level of efficacy and protection, this vaccine was administered transdermally and tested in a pre-clinical mouse model. The M2e VLP was encapsulated into a polymeric matrix with the addition of Alhydrogel® and Monophosphoryl Lipid-A (MPL-A®), together referred to as AS04. The M2e VLP formulations induced IgG titers, with increased levels of IgG1 in the M2e VLP MP groups and further elevated levels of IgG2a were found specifically in the M2e VLP MP Adjuvant group. This trend in humoral immunity was also observed from a cell-mediated standpoint, where M2e VLP MP groups showed increased expression in CD4+ T cells in the spleen and the lymph node and high levels of CD8+ T cells in the lymph node. Taken together, the results illustrate the immunogenic potential of the matrix-2 protein virus-like particle (M2e VLP) vaccine.

scholarly journals Live cell PNA labelling enables erasable fluorescence imaging of membrane proteins

2020 ◽  
Author(s):  
Georgina C. Gavins ◽  
Katharina Gröger ◽  
Michael D. Bartoschek ◽  
Philipp Wolf ◽  
Annette G. Beck-Sickinger ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Cho Cells ◽  
Fluorescent Dyes ◽  
Coiled Coil ◽  
Dna Nanotechnology ◽  
Nucleic Acid Hybridization ◽  
Landing Platform ◽  
Acid Protein ◽  
Reaction Proceeds ◽  
A Cell

AbstractDNA nanotechnology is an emerging field, which promises fascinating opportunities for the manipulation and imaging of proteins on a cell surface. The key to progress in the area is the ability to create the nucleic acid-protein junction in the context of living cells. Here we report a covalent labelling reaction, which installs a biostable peptide nucleic acid (PNA) tag. The reaction proceeds within minutes and is specific for proteins carrying a 2 kDa coiled coil peptide tag. Once installed the PNA label serves as a generic landing platform that enables the recruitment of fluorescent dyes via nucleic acid hybridization. We demonstrate the versatility of this approach by recruiting different fluorophores, assembling multiple fluorophores for increased brightness, and achieving reversible labelling by way of toehold mediated strand displacement. Additionally, we show that labelling can be carried out using two different coiled coil systems, with EGFR and ETBR, on both HEK293 and CHO cells. Finally, we apply the method to monitor internalization of EGFR on CHO cells.

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