Escherichia coli signal peptidase recognizes and cleaves the signal sequence of xylanase from a newly isolated Bacillus subtilis strain R5

2011 ◽  
Vol 76 (3) ◽  
pp. 347-349 ◽  
Author(s):  
A. Jalal ◽  
N. Rashid ◽  
N. Ahmed ◽  
S. Iftikhar ◽  
M. Akhtar
Keyword(s):  
Escherichia Coli ◽  
Bacillus Subtilis ◽  
Signal Sequence ◽  
Signal Peptidase ◽  
Subtilis Strain ◽  
Bacillus Subtilis Strain R5

scholarly journals Characterization of the Bacillus subtilis ywtD Gene, Whose Product Is Involved in γ-Polyglutamic Acid Degradation

2003 ◽  
Vol 185 (7) ◽  
pp. 2379-2382 ◽  
Author(s):  
Takao Suzuki ◽  
Yasutaka Tahara
Keyword(s):  
Escherichia Coli ◽  
Bacillus Subtilis ◽  
Glutamic Acid ◽  
Recombinant Plasmid ◽  
Signal Sequence ◽  
High Molecular Mass ◽  
Enzymatic Analysis ◽  
Polyglutamic Acid ◽  
Partial Similarity

ABSTRACT The ywtD gene, which codes for an enzyme that degrades γ-polyglutamic acid (PGA), was cloned from Bacillus subtilis IFO16449. The gene is located immediately downstream of ywsC and ywtABC, a PGA operon involved in PGA biosynthesis, and it showed partial similarity to genes coding for dl-endopeptidase, a peptidoglycan-degrading enzyme. The ywtD gene, from which signal sequence is excised, was inserted into pET15b, and the recombinant plasmid was then transformed into Escherichia coli. Histidine-tagged YwtD was purified from sonicated cells of the transformant. The purified YwtD degraded PGA to yield two hydrolyzed products, a high-molecular-mass product (490 kDa with nearly 100% l-glutamic acid) and an 11-kDa product (with d-glutamic acid and l-glutamic acid in an 80:20 ratio). This finding and results of enzymatic analysis of the two products with carboxypeptidase G suggest that YwtD is a novel enzyme cleaving the γ-glutamyl bond only between d- and l-glutamic acids of PGA, and it may be designated γ-dl-glutamyl hydrolase.

Gene cloning and characterization of glycine oxidase from newly isolated Bacillus subtilis strain R5

Biologia ◽  
2011 ◽  
Vol 66 (1) ◽  
pp. 1-7 ◽  
Author(s):  
Farrukh Jamil ◽  
Naeem Rashid ◽  
Qurra-tul-Ann Gardner ◽  
Muhammad Akhtar
Keyword(s):  
Enzyme Activity ◽  
Bacillus Subtilis ◽  
Specific Activity ◽  
Subtilis Strain ◽  
Amino Acid Residues ◽  
Expression And Purification ◽  
Gene Encoding ◽  
Glycine Oxidase ◽  
Bacillus Subtilis Strain R5

AbstractThe gene encoding the glycine oxidase from Bacillus subtilis strain R5 (goxR) was cloned and expressed in Escherichia coli. The gene consisted of 1,110 nucleotides that encoded a protein (GoxR) of 369 amino acid residues with a molecular mass of 40,761 Da. The GoxR exhibited 98.6% identity with glycine oxidase from B. subtilis strain 168. Gene expression and purification of the recombinant GoxR were performed. The recombinant GoxR existed in a homotetramer form. The recombinant protein effectively catalyzed the oxidation of glycine and d-alanine. The specific activity of the purified recombinant GoxR was 0.96 U/mg when glycine was used as a substrate and 1.0 U/mg when d-alanine was substrate. The enzyme displayed its highest activity at pH 8.0 and at a temperature of 50°C. The activation energy of the reaction catalyzed by the enzyme was calculated to be 26 kJ/mol. The enzyme activity was significantly inhibited in the presence of organic solvents. No enhancement of enzyme activity was observed in the presence of metal cations. The experimental results presented in this study demonstrate that the enzyme was a bonafide glycine oxidase.

scholarly journals Processing of chimeric mammalian cytochrome b5 precursors in Escherichia coli: reaction specificity of signal peptidase and identification of an aminopeptidase in post-translocational processing

1993 ◽  
Vol 293 (3) ◽  
pp. 751-756 ◽  
Author(s):  
V Harding ◽  
A Karim ◽  
N Kaderbhai ◽  
A Jones ◽  
A Evans ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Alkaline Phosphatase ◽  
Signal Sequence ◽  
Cytochrome B5 ◽  
Arginine Residue ◽  
Signal Peptidase ◽  
Apparent Absence ◽  
Sequence Deletion ◽  
Reaction Specificity

A chimeric precursor interlinked by an arginine residue between the full-length signal sequence of alkaline phosphatase and the eukaryotic cytoplasmic cytochrome b5 was constructed. Expression of the chimeric precursor protein in Escherichia coli resulted in efficient export of spectrally authentic cytochrome b5 into the periplasm [Karim, Harding, Evans, Kaderbhai and Kaderbhai (1993) Bio/Technology 11, 612-618]. On sequencing, the apparent absence of arginine at the N-terminus of the secreted cytochrome b5 implied that the chimera was either miscleaved by signal peptidase or further processed following signal excision by an uncharacterized peptidase. The influence of the N-terminal region of cytochrome b5 on the unusual processing of the chimeric precursor was investigated by engineering a number of variant forms in which the region between Arg+1 and the mature portion of cytochrome b5 was extended and varied. Observations of the in vivo processed patterns of these variant cytochrome b5 forms exported into the periplasm revealed that the absence of arginine was due to neither miscleavage of the translocated precursor by the signal peptidase nor the nature of the early region of cytochrome b5. In fact, the selective excision of the arginine residue occurred subsequent to signal sequence deletion by an aminopeptidase which was sensitive to the metal chelator o-phenanthroline. We show that this aminopeptidase also participates in the trimming of the N-terminal arginine residue of the bacterial alkaline phosphatase to generate the three isoenzymes in the periplasm.

scholarly journals A Polysaccharide Deacetylase Homologue, PdaA, in Bacillus subtilis Acts as an N-Acetylmuramic Acid Deacetylase In Vitro

2005 ◽  
Vol 187 (4) ◽  
pp. 1287-1292 ◽  
Author(s):  
Tatsuya Fukushima ◽  
Toshihiko Kitajima ◽  
Junichi Sekiguchi
Keyword(s):  
Mass Spectrometry ◽  
Escherichia Coli ◽  
High Performance Liquid Chromatography ◽  
Bacillus Subtilis ◽  
Liquid Chromatography ◽  
High Performance ◽  
Signal Sequence ◽  
Reverse Phase

ABSTRACT A polysaccharide deacetylase homologue, PdaA, was determined to act as an N-acetylmuramic acid deacetylase in vitro. Histidine-tagged truncated PdaA (with the putative signal sequence removed) was overexpressed in Escherichia coli cells and purified. Measurement of deacetylase activity showed that PdaA could deacetylate peptidoglycan treated with N-acetylmuramoyl-l-alanine amidase CwlH but could not deacetylate peptidoglycan treated with or without dl-endopeptidase LytF (CwlE). Reverse-phase high-performance liquid chromatography and mass spectrometry (MS) and MS-MS analyses indicated that PdaA could deacetylate the N-acetylmuramic acid residues of purified glycan strands derived from Bacillus subtilis peptidoglycan.

scholarly journals Non-functional expression of Escherichia coli signal peptidase I in Bacillus subtilis

1991 ◽  
Vol 137 (9) ◽  
pp. 2073-2083 ◽  
Author(s):  
J. M. van Dijl ◽  
A. de Jong ◽  
H. Smith ◽  
S. Bron ◽  
G. Venema
Keyword(s):  
Escherichia Coli ◽  
Bacillus Subtilis ◽  
Functional Expression ◽  
Signal Peptidase ◽  
Signal Peptidase I
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