Posttranslational processing of the xylanase Xys1L from Streptomyces halstedii JM8 is carried out by secreted serine proteases

• Published in: Microbiology (Society for General Microbiology) 2003-07, Vol.149 (7), p.1623-1632
• Main Authors: Fernandez-Abalos, Jose M, Reviejo, Veronica, Diaz, Margarita, Rodriguez, Sonia, Leal, Fernando, Santamaria, Ramon I
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Bacterial Proteins; Bacteriology; Base Sequence; Biological and medical sciences; Cloning, Molecular; DNA, Bacterial - genetics; Fundamental and applied biological sciences. Psychology; Gene Expression; Genes, Bacterial; Genetics; Metabolism. Enzymes; Microbiology; Molecular Sequence Data; Protein Processing, Post-Translational; Serine Endopeptidases - genetics; Serine Endopeptidases - metabolism; Serine Proteinase Inhibitors - genetics; Serine Proteinase Inhibitors - metabolism; Serine Proteinase Inhibitors - pharmacology; SLPI protein; SpB gene; Streptomyces - enzymology; Streptomyces - genetics; Streptomyces halstedii; Streptomyces lividans; Xylosidases - genetics; Xylosidases - metabolism; Xys1L protein; xysA gene
• ISSN: 1350-0872; 1465-2080
• DOI: 10.1099/mic.0.26113-0

Instituto de Microbiología Bioquímica/Departamento de Microbiología y Genética, Consejo Superior de Investigaciones Científicas (CSIC)/Universidad de Salamanca, Edificio Departamental, Campus Miguel de Unamuno, 37007 Salamanca, Spain Correspondence Ramón I. Santamaría santa{at}usal.es The xylanase Xys1L from Streptomyces halstedii JM8 is known to be processed extracellularly, to produce a protein of 33·7 kDa, Xys1S, that retains catalytic activity but not its cellulose-binding capacity. This paper demonstrates that at least five serine proteases isolated from Streptomyces spp. have the ability to process the xylanase Xys1L. The genes of two of these extracellular serine proteases, denominated SpB and SpC, were cloned from Streptomyces lividans 66 (a strain commonly used as a host for protein secretion), sequenced, and overexpressed in S. lividans ; both purified proteases were able to process Xys1L in vitro . Three other previously reported purified Streptomyces serine proteases, SAM-P20, SAM-P26 and SAM-P45, also processed Xys1L in vitro . The involvement of serine proteases in xylanase processing-degradation in vivo was demonstrated by co-expression of the xylanase gene ( xysA ) and the gene encoding the serine protease inhibitor (SLPI) from S. lividans . Co-expression prevented processing and degradation of Xys1L and resulted in a threefold increase in the xylanase activity present in the culture supernatant. SpB and SpC also have the capacity to process other secreted proteins such as p40, a cellulose-binding protein from S. halstedii JM8, but do not have any clear effect on other secreted proteins such as amylase (Amy) from Streptomyces griseus and xylanase Xyl30 from Streptomyces avermitilis . Abbreviations: SLPI, Streptomyces lividans (subtilisin-like) serine protease inhibitor; STI, soybean trypsin inhibitor; TLCK, N - p -tosyl- L -lysine chloromethyl ketone hydrochloride These two authors contributed equally to the results obtained in this paper. The GenBank accession number for the sequences determined in this work are AJ496191 and AJ496192 .