title: Extracellular serine proteases from Stenotrophomonas maltophilia: Screening, isolation and heterologous expression in E. coli.
abstract: A large strain collection comprising antagonistic bacteria was screened for novel detergent proteases. Several strains displayed protease activity on agar plates containing skim milk but were inactive in liquid media. Encapsulation of cells in alginate beads induced protease production. Stenotrophomonas maltophilia emerged as best performer under washing conditions. For identification of wash-active proteases, four extracellular serine proteases called StmPr1, StmPr2, StmPr3 and StmPr4 were cloned. StmPr2 and StmPr4 were sufficiently overexpressed in E. coli. Expression of StmPr1 and StmPr3 resulted in unprocessed, insoluble protein. Truncation of most of the C-terminal domain which has been identified by enzyme modeling succeeded in expression of soluble, active StmPr1 but failed in case of StmPr3. From laundry application tests StmPr2 turned out to be a highly wash-active protease at 45°C. Specific activity of StmPr2 determined with suc-L-Ala-L-Ala-L-Pro-l-Phe-p-nitroanilide as the substrate was 17±2U/mg. In addition we determined the kinetic parameters and cleavage preferences of protease StmPr2.
authors: ACIB GmbH, Petersgasse 14, 8010 Graz, Austria. doris.ribitsch@acib.at
authors: D
authors: D
authors: Ribitsch
authors: S
authors: S
authors: Heumann
authors: W
authors: W
authors: Karl
authors: J
authors: J
authors: Gerlach
authors: R
authors: R
authors: Leber
authors: R
authors: R
authors: Birner-Gruenberger
authors: K
authors: K
authors: Gruber
authors: I
authors: I
authors: Eiteljoerg
authors: P
authors: P
authors: Remler
authors: P
authors: P
authors: Siegert
authors: J
authors: J
authors: Lange
authors: K H
authors: KH
authors: Maurer
authors: G
authors: G
authors: Berg
authors: G M
authors: GM
authors: Guebitz
authors: H
authors: H
authors: Schwab
mesh_terms: D000464:Alginates / Q000737:chemistry; D000818:Animals; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D003470:Culture Media / Q000378:metabolism; D003902:Detergents / Q000737:chemistry; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005110:Extracellular Space / Q000378:metabolism; D020723:Glucuronic Acid / Q000737:chemistry; D006603:Hexuronic Acids / Q000737:chemistry; D007328:Insulin / Q000378:metabolism; D008892:Milk / Q000378:metabolism; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D057057:Serine Proteases / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D020615:Stenotrophomonas maltophilia / Q000201:enzymology* / Q000235:genetics
chemical_list: D000464:Alginates; D001426:Bacterial Proteins; D003470:Culture Media; D003902:Detergents; D006603:Hexuronic Acids; D007328:Insulin; D011994:Recombinant Proteins; D020723:Glucuronic Acid; D057057:Serine Proteases