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PMID: 7751281 Published · ppublish English Comparative Study Journal Article Research Support, Non-U.S. Gov't

Purification and characterization of the phospho-alpha(1,1)glucosidase (TreA) of Bacillus subtilis 168.

Journal of bacteriology ·Vol. 177 ·No. 10 ·1995-05-00 ·Pages 2721-6

Gotsche S, Dahl MK

Abstract

The intracellular phospho-alpha(1,1)glucosidase TreA from Bacillus subtilis has been overproduced in Escherichia coli and purified by ion-exchange chromatography and gel filtration. The molecular mass, estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, was 64 kDa. Isoelectric focusing indicated homogeneity of the protein, and its pI was determined to be 4.3. Characterization of the enzyme showed a protein which is stable up to 44 degrees C after temperature treatment for 15 min. The temperature optimum was found to be 37 degrees C, and the pH optimum was 4.5. TreA activity is stimulated by high salt concentrations with different efficiencies depending on the kind of salt. When increasing amounts of ammonium sulfate are used, the increase of TreA activity is correlated with a conformational change of the protein or dimerization. The substrate specificity of the purified enzyme was characterized, showing additionally that trehalose is also hydrolyzed, but to a much smaller extent than trehalose-6-phosphate. In vitro, the presence of glucose reduces TreA activity, indicating product inhibition of the enzyme.

Related Genes
treA
MeSH Terms
Bacillus subtilis/enzymology,genetics Disaccharidases/genetics,isolation & purification,metabolism Enzyme Stability Escherichia coli/genetics Fructose/metabolism Glucose/metabolism Isoelectric Focusing Molecular Weight Potassium Chloride/pharmacology Recombinant Proteins/isolation & purification,metabolism Sodium Chloride/pharmacology Substrate Specificity Sugar Phosphates/metabolism Trehalose/analogs & derivatives,metabolism
Chemicals
Recombinant Proteins Sugar Phosphates Fructose trehalose-6-phosphate Sodium Chloride Potassium Chloride Trehalose Disaccharidases alpha, alpha-phosphotrehalase Glucose
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Gotsche S
Lehrstuhl für Mikrobiologie, Friedrich-Alexander Universität Erlangen-Nürnberg, Federal Republic of Germany.
Dahl M K
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Article Info
Journal
Journal of bacteriology
Abbr.
J Bacteriol
ISSN
0021-9193
DOI
10.1128/jb.177.10.2721-2726.1995
Published
1995-05-00
Pages
2721-6
Language
English
Region
United States
NLM ID
2985120R
PMCID
PMC176942
Subset
IM
External Links
PubMed source DOI: 10.1128/jb.177.10.2721-2726.1995 PMC Full Text View journal details
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