Purification and partial characterization of an extracellular serine‐proteinase of Streptomyces cyaneus isolated from Brazilian cerrado soil

Streptomyces cyaneus, a micro‐organism isolated from Brazilian cerrado soil, produces an extracellular proteinase (SCP), which was purified 22‐fold to homogeneity from culture supernatant fluid, using a single aprotinin‐agarose affinity chromatography step. It is produced at a level corresponding to...

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Bibliographic Details
Published in:	Journal of applied microbiology Vol. 87; no. 4; pp. 557 - 563
Main Authors:	Petinate, S.D.G, Branquinha, M.H, Coelho, R.R.R, And, A.B. Vermelho, Giovanni‐De‐Simone, S
Format:	Journal Article
Language:	English
Published:	Oxford, UK Blackwell Science Ltd 01-10-1999 Blackwell Science
Subjects:	Biological and medical sciences Biology of microorganisms of confirmed or potential industrial interest Biotechnology Brazil cerrado soils Chromatography, Gel Electrophoresis, Polyacrylamide Gel Fundamental and applied biological sciences. Psychology Hydrogen-Ion Concentration Miscellaneous Mission oriented research Molecular Weight Serine Endopeptidases - chemistry Serine Endopeptidases - isolation & purification Serine Endopeptidases - metabolism serine proteinase Serine Proteinase Inhibitors - pharmacology serine proteinases soil bacteria Soil Microbiology Streptomyces - enzymology Streptomyces cyaneus Temperature Tosyl Compounds - pharmacology Tosylphenylalanyl Chloromethyl Ketone - pharmacology Brazil Actinomycetales Peptidases Streptomycetaceae Purification Serine endopeptidases Enzyme Bacteria Hydrolases Actinomycetes Kinetic parameter Physicochemical properties
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Summary:	Streptomyces cyaneus, a micro‐organism isolated from Brazilian cerrado soil, produces an extracellular proteinase (SCP), which was purified 22‐fold to homogeneity from culture supernatant fluid, using a single aprotinin‐agarose affinity chromatography step. It is produced at a level corresponding to approximately 15% of total protein, but its physiological function has yet to be determined. The molecular mass of this S. cyaneus proteinase was estimated to be 120 kDa by gel filtration high performance liquid chromatography, and it migrates by SDS‐PAGE as a single band of 30 kDa. It was optimally active at 25 °C and pH 9·0, and was fully inhibited by the serine‐proteinase inhibitors PMSF and TPCK. A Km value of 1·86 × 10⁻⁵ mmol l⁻¹, and Vmax of 2·0 × 10⁻² mmol l⁻¹ (Abs₂₄₇ nmμg⁻¹ min⁻¹), were calculated for α‐N‐p‐tosyl‐ l‐arginine‐methyl ester (TAME) as substrate.
Bibliography:	http://dx.doi.org/10.1046/j.1365-2672.1999.00852.x ObjectType-Article-2 SourceType-Scholarly Journals-1 ObjectType-Feature-1 content type line 23 ObjectType-Article-1 ObjectType-Feature-2
ISSN:	1364-5072 1365-2672
DOI:	10.1046/j.1365-2672.1999.00852.x