Secretion by overexpression and purification of the water-soluble Streptomyces K15 DD-transpeptidase/penicillin-binding protein

Though synthesized with a cleavable signal peptide and devoid of membrane anchors, the 262-amino-acid-residue Streptomyces K15 DD-transpeptidase/penicillin-binding protein is membrane-bound. Overexpression in Streptomyces lividans resulted in the export of an appreciable amount of the synthesized pr...

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Bibliographic Details
Published in:	Biochemical journal Vol. 288; no. 1; pp. 87 - 91
Main Authors:	PALOMEQUE-MESSIA, P, QUITTRE, V, LEYH-BOUILLE, M, NGUYEN-DISTECHE, M, GERSHATER, C. J. L, DACEY, I. K, DUSART, J, VAN BEEUMEN, J, GHUYSEN, J.-M
Format:	Journal Article Web Resource
Language:	English
Published:	Colchester Portland Press 15-11-1992
Subjects:	Amino Acid Sequence Bacterial Proteins Bacteriology Biochemistry, biophysics & molecular biology Biochimie, biophysique & biologie moléculaire Biological and medical sciences Carrier Proteins - genetics Carrier Proteins - isolation & purification Carrier Proteins - metabolism Cell Membrane - enzymology characterization Enzyme Stability Fundamental and applied biological sciences. Psychology Gene Expression Hexosyltransferases Hot Temperature Life sciences Metabolism. Enzymes Microbiology Molecular Sequence Data Molecular Weight Muramoylpentapeptide Carboxypeptidase - genetics Muramoylpentapeptide Carboxypeptidase - isolation & purification Muramoylpentapeptide Carboxypeptidase - metabolism Penicillin-Binding Proteins Penicillins - metabolism Peptidyl Transferases Plasmids purification Sciences du vivant serine-type D-Ala-D-Ala carboxypeptidase Sodium Chloride Solubility Streptomyces - enzymology Streptomyces - genetics Streptomyces lividans thermal stability Transformation, Bacterial Actinomycetales Streptomycetaceae Purification Enzymatic activity Enzyme Secretion Streptomyces lividans Gene overexpression Bacteria Actinomycetes Physicochemical properties Penicillin binding protein
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Summary:	Though synthesized with a cleavable signal peptide and devoid of membrane anchors, the 262-amino-acid-residue Streptomyces K15 DD-transpeptidase/penicillin-binding protein is membrane-bound. Overexpression in Streptomyces lividans resulted in the export of an appreciable amount of the synthesized protein (4 mg/litre of culture supernatant). The water-soluble enzyme was purified close to protein homogeneity with a yield of 75%. It requires the presence of 0.5 M-NaCl to remain soluble. It is indistinguishable from the detergent-extract wild-type enzyme with respect to molecular mass, thermostability, transpeptidase activity and penicillin-binding capacity.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 scopus-id:2-s2.0-0026474765
ISSN:	0264-6021 1470-8728 1470-8728
DOI:	10.1042/bj2880087