Amino sugars in the glycoprotein toxin from Bacillus thuringiensis subsp. israelensis

Pfannenstiel, Mary Ann; Muthukumar, Ganapathy; Couche, Graham A.; Nickerson, Kenneth W.

doi:10.1128/jb.169.2.796-801.1987

Cited by 28 publications

(22 citation statements)

References 31 publications

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“…Pfannenstiel et al (27) have reported that B. thuringiensis subsp. israelensis crystal proteins contain amino sugars.…”

Section: Methodsmentioning

confidence: 99%

“…Electrophoresis of the solubilized inclusions reveals four major polypeptides with molecular masses of 135, 128, 67, and 27 kilodaltons (kDa) (2,35). These polypeptides are glycosylated (27) and are mosquitocidal; the 27-kDa peptide is also hemolytic and cytolytic (32). Many studies have also reported the presence of several minor crystal components: a 230-kDa species (probably an aggregate of the 135-or 128-kDa peptide or both; 38), a toxic 58-kDa polypeptide (derived from a 72-kDa peptide; 33), a 54-kDa peptide (probably a dimer of the 27-kDa peptide; 7), and a nontoxic 38-kDa peptide (probably a breakdown product of the 67-kDa peptide; 7,19).…”

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confidence: 99%

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A 20-kilodalton protein is required for efficient production of the Bacillus thuringiensis subsp. israelensis 27-kilodalton crystal protein in Escherichia coli

1989

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The 27-kilodalton (kDa) mosquitocidal protein gene from Bacillus thuringiensis subsp. israelensis has been cloned as a 10-kilobase (kb) HindIII fragment from plasmid DNA; efficient expression in Escherichia coli KM1 depends on a region of DNA located approximately 4 kb upstream (K. McLean and H. R. Whiteley, J. Bacteriol. 169:1017-1023, 1987). We have cloned the upstream DNA region and show that it contains a complete open reading frame (ORF) encoding a protein with a molecular mass of 19,584 Da. Sequencing of adjacent stretches of DNA revealed two partial ORFs: one has 55.2% identity in an overlap of 319 amino acids to the putative transposase of IS231 of B. thuringiensis subsp. thuringiensis, and the other, a 78-codon partial ORF, may be the carboxyl terminus of the 67-kDa protein previously observed in maxicells of strain KM1. A 0.8-kb fragment containing only the 20-kDa protein gene greatly enhanced the expression of the 27-kDa protein in E. coli. The introduction of nonsense codons into the 20-kDa protein gene ORF abolished this effect, indicating that the gene product, not the mRNA or DNA, is required for the enhancement. The effect of the 20-kDa protein gene on various fusions of lacZ to the 27-kDa protein gene suggests that the 20-kDa protein acts after the initiation of translation of the 27-kDa protein gene.

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“…Pfannenstiel et al (27) have reported that B. thuringiensis subsp. israelensis crystal proteins contain amino sugars.…”

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

A 20-kilodalton protein is required for efficient production of the Bacillus thuringiensis subsp. israelensis 27-kilodalton crystal protein in Escherichia coli

1989

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“…In Gram-positive bacteria, GalN and glucosamine (GlcN) have been reported in the glycosylation motif of a glycoprotein toxin from Bacillus thuringiensis subsp. israelensis and proposed to be important to the biological activities of the toxin (10). GalN was also identified as a possible component of a galactose-containing polysaccharide from Streptococcus mutans (11).…”

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confidence: 99%

Biosynthetic Origin of the Galactosamine Substituent of Arabinogalactan in Mycobacterium tuberculosis

Škovierová

Larrouy-Maumus

Phạm

et al. 2010

Journal of Biological Chemistry

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The arabinogalactan (AG) of slow growing pathogenic Mycobacterium spp. is characterized by the presence of galactosamine (GalN) modifying some of the interior branched arabinosyl residues. The biosynthetic origin of this substituent and its role(s) in the physiology and/or pathogenicity of mycobacteria are not known. We report on the discovery of a polyprenylphospho-N-acetylgalactosaminyl synthase (PpgS) and the glycosyltransferase Rv3779 from Mycobacterium tuberculosis required, respectively, for providing and transferring the GalN substrate for the modification of AG. Disruption of either ppgS (Rv3631) or Rv3779 totally abolished the synthesis of the GalN substituent of AG in M. tuberculosis H37Rv. Conversely, expression of ppgS in Mycobacterium smegmatis conferred upon this species otherwise devoid of ppgS ortholog and any detectable polyprenyl-phospho-N-acetylgalactosaminyl synthase activity the ability to synthesize polyprenyl-phospho-N-acetylgalactosamine (polyprenyl-P-GalNAc) from polyprenyl-P and UDP-GalNAc. Interestingly, this catalytic activity was increased 40 -50-fold by co-expressing Rv3632, the encoding gene of a small membrane protein apparently co-transcribed with ppgS in M. tuberculosis H37Rv. The discovery of this novel lipid-linked sugar donor and the involvement of a the glycosyltransferase C-type glycosyltransferase in its transfer onto its final acceptor suggest that pathogenic mycobacteria modify AG on the periplasmic side of the plasma membrane.The availability of a ppgS knock-out mutant of M. tuberculosis provides unique opportunities to investigate the physiological function of the GalN substituent and the potential impact it may have on host-pathogen interactions.Galactosamine (D-GalN) was recognized as a minor covalently bound amino sugar component of the cell wall of slow growing mycobacteria (Mycobacterium tuberculosis H37Ra, Mycobacterium lepraemurium, Mycobacterium microti) as early as in the 1970s (1-3). Advances in mass spectrometry later coupled with the use of an arabinanase isolated from Mycobacterium smegmatis allowed the GalN substituent to be mapped to the arabinogalactan (AG) 7 component of the cell wall (3), and more specifically to the C2 position of a portion of the internal 3,5-branched D-Araf residues in the AG of M. tuberculosis (4, 5). D-GalN was estimated to occur at the level of about one residue per entire AG molecule (3-5). Interestingly, a similar sugar residue was found to substitute the AG of Mycobacterium avium, Mycobacterium kansasii, Mycobacterium bovis BCG (3) and Mycobacterium leprae, 8 but not that of M. smegmatis, M. neoaurum, and M. phlei (3-5), suggesting that fast growing Mycobacterium spp. are devoid of GalN substituent.The function and biosynthetic origin of this substituent are not known. Non-N-acetylated hexosamine residues, particularly GalN residues, are rather uncommon in prokaryotes. Thus far, they essentially have been described as components of the lipopolysaccharide (LPS) of some Gram-negative bacteria (6 -9). In Francisella tularensis a...

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“…Such proteins are speculated to account for the existence of the enzymatic rather than nonenzymatic type of glycosylation in prokaryotes (27). The only bacterial glycoprotein considered so far as being a result of glycation is the Bacillus thuringiensis crystal protein (28,29). However, because this protein is secreted in the culture medium, it was concluded that the "sugars detected were the product of fermentation conditions rather than bacterial genetics" (30).…”

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Glycation and Post-translational Processing of Human Interferon-γ Expressed in Escherichia coli

Mironova

Niwa

Dimitrova

et al. 2003

Journal of Biological Chemistry

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Amino sugars in the glycoprotein toxin from Bacillus thuringiensis subsp. israelensis

Abstract: The carbohydrate content of purified Bacillus

Cited by 28 publications

References 31 publications

A 20-kilodalton protein is required for efficient production of the Bacillus thuringiensis subsp. israelensis 27-kilodalton crystal protein in Escherichia coli

A 20-kilodalton protein is required for efficient production of the Bacillus thuringiensis subsp. israelensis 27-kilodalton crystal protein in Escherichia coli

Biosynthetic Origin of the Galactosamine Substituent of Arabinogalactan in Mycobacterium tuberculosis

Glycation and Post-translational Processing of Human Interferon-γ Expressed in Escherichia coli

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