Abstract
Production of proteins in secretary form is one of the important factors affecting fermentation. The Tat (twin arginine translocation) protein secretion system, which includes the proteins TatA, TatB, and TatC, was identified in the genomic sequence of Streptomyces griseus IFO13350. The tatA and tatC genes were organized into a polycistronic operon, whereas tatB was located separately on the chromosome. Comparison of amino acid sequences suggested that TatC was a membrane-spanning protein, whereas TatA and TatB were found to be cytoplasmic proteins. Analysis of extracellular proteins and N-terminal amino acid sequencing revealed that secretion of SGR5556 was significantly enhanced by overexpression of TatAC in S. griseus HH1. Further, enzymatic study showed that SGR5556 encoded a glycerophosphoryl diester phosphodiesterase. In addition, other hydrolase activities, such as those of amylase, total protease, metalloprotease, trypsin, chymotrypsin, and Leuaminopeptidase, were also enhanced by 3, 3, 2.6, 2.3, 5.4, and 2.5 fold, respectively, in S. griseus upon TatAC overexpression. Overexpression of TatAC induced the production of a greenish-yellow pigment in S. griseus HH1 as well as more abundant sporulation at an earlier stage in Streptomyces coelicolor A3(2). In silico analysis by TatFIND, SignalP, and TMHMM identified 19 binding proteins, 28 enzymatic proteins, and 27 other proteins with unknown functions as putative TatAC-dependent secretary proteins. These results clearly indicate that TatA and TatC constitute a functional Tat system in S. griseus. Additionally, the S. griseus Tat system can be useful for the production of valuable proteins, including many hydrolytic enzymes and candidates of Tat-dependent secretary proteins, under industrial conditions.
Similar content being viewed by others
References
Pusley, A. (1993) The complete general secretory pathway in Gram-negative bacteria. Microbiol. Rev. 57: 50–108.
Santini, C. L., B. Ize, A. Chanal, M. Müller, G. Giordano, and L. F. Wu (1998) A novel sec-independent periplasmic protein translocation pathway in Escherichia coli. EMBO J. 17: 101–121.
Yen, M. R., Y. H. Tseng, E. H. Nguyen, L. F. Wu, and M. H. Saier Jr (2002) Sequence and phylogenetic analyses of the twin-arginine targeting (Tat) protein export system. Arch. Microbiol. 177: 441–450.
Berks, B. C., F. Sargent, and T. Palmer (2000) The Tat protein export pathway. Mol. Microbiol. 35: 260–274.
Bogsch, E. G., F. Sargent, N. R. Stanley, B. C. Berks, C. Robinson, and T. Palmer (1998) An essential component of a novel bacterial protein export system with homologues in plastids and mitochondria. J. Biol. Chem. 273: 18003–18006.
Jongbloed J. D., U. Grieger, H. Antelmann, M. Hecker, R. Nijland, S. Bron, and J. M. van Diji (2004) Two minimal Tat translocases in Bacillus. Mol. Microbiol. 54: 1319–1325.
Schaerlaekens, K., L. Van Mellaert, E. Lammertyn, N. Geukens, and J. Anné (2004) The importance of the Tat-dependent protein secretion pathway in Streptomyces as revealed by phenotypic changes in tat deletion mutants and genome analysis. Microbiol. 150: 21–31.
Kikuchi, Y., H. Itaya, M. Date, K. Matsui, and L. F. Wu (2009) TatABC overexpression improves Corynebacterium glutamicum Tat-dependent protein secretion. Appl. Environ. Microbiol. 75: 603–607.
McDonough, J. A., K. E. Hacker, A. R. Flores, M. S. Pavelka Jr., and M. Braunstein (2005) The twin-arginine translocation pathway of Mycobacterium smegmatis is functional and required for the export of mycobacterial beta-lactamases. J. Bacteriol. 187: 7677–7679.
Shin, J. H., J. H. Choi, O. -S. Lee, Y. -M. Kim, D. -S. Lee, Y. Y. Kwak, W. C. Kim, and I. K. Rhee (2009) Thermostable xylanase from Streptomyces thermocyaneoviolaceus for optimal production of xylooligosaccharides. Biotechnol. Bioproc. Eng. 14: 391–399.
Vujaklija, D., K. Ueda, S. -K. Hong, T. Beppu, and S. Horinouchi (1991) Identification of an A-factor-dependent promoter in the streptomycin biosynthetic gene cluster of Streptomyces griseus. Mol. Gen. Genet. 229: 119–128.
Hwang, C. K., H. S. Kim, Y. S. Hong, S. K. Hong, and J. J. Lee (1995) Expression of Streptomyces peucetius for doxorubicin resistance and aklavinone 11-hydroxylase in Streptomyces galilaeus ATCC 31133 and production of hybrid aclacinomycin. Antimicrob. agent Chemother. 39: 1616–1620.
Binnie, C., J. D. Cossar, and D. I. Stewart (1997) Heterologous biopharmaceutical protein expression in Streptomyces. Trends. Biotechnol. 15: 315–320.
Lammertyn, E., L. Van Mellaert, S. Schacht, C. Dillen, E. Sablon, A. Van Broekhoven, and J. Anné (1997) Evaluation of a novel subtilisin inhibitor gene and mutant derivatives for the expression and secretion of mouse tumor necrosis factor alpha by Streptomces lividans. Appl. Environ. Microbiol. 63: 1808–1813.
Kato, J. Y., W. J. Chi, Y. Ohnishi, S. K. Hong, and S. Horinouchi (2005) Transcriptional control by A-factor of two trypsin genes in Streptomyces griseus. J. Bacteriol. 187: 286–295.
Tomono, A., Y. Tasi, Y. Ohnishi, and S. Horinouchi (2005) Three chymotrypsin genes are members of the AdpA regulon in the A-factor regulatory cascade in Streptomyces griseus. J. Bacteriol. 187: 6341–6353.
Choi, E. Y., E. A. Oh, J. H. Kim, D. K. Kang, and S. K. Hong (2007) Distinct regulation of the sprC gene encoding Streptomyces griseus protease C from other chymotrypsin genes in Streptomyces griseus IFO13350. J. Microbiol. Biotechnol. 17: 81–88.
Ohnishi, Y., J. Ishikawa, H. Hara, H. Suzuki, M. Ikenoya, H. Ikeda, A. Yamashita, M. Hattori, and S. Horinouchi (2008) Genome sequence of the streptomycin-producing microorganism Streptomyces griseus IFO13350. J. Bacteriol. 190: 4050–4060.
Kieser, H., M. J. Bibb, M. J. Buttner, K. F. Chater, and D. A. Hopwood (2000) Practical Streptomyces genetics. The John Innes Foundation, Norwich, UK.
Vara, J., M. Lewandowska-Skarbek, Y. G. Wang, S. Donadio, and C. R. Hutchinson (1989) Cloning of genes governing the deoxysugar portion of the erythromycin biosynthesis pathway in Saccharopolyspora erythraea (Streptomyces erythreus). J. Bacteriol. 171: 5872–5881.
Sambrook, J. and D. W. Russell (2001) Molecular cloning: A laboratory manual. 3rd ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA.
Laemmli, U. K. (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680–685.
Bradford, M. M. (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: 248–254.
Kim, J. M. and S. K. Hong (2000) Streptomyces griseus HH1, an A-factor deficient mutant, produces diminished level of trypsin and increased level of metalloproteases. J. Microbiol. 38: 160–168.
Rose, R. W., T. Bruser, J. C. Kissinger, and M. Pohlschröder (2002) Adaptation of protein secretion to extremely high-salt conditions by extensive use of the twin-arginine translocation pathway. Mol. Microbiol. 45: 943–950.
Larson, T. J., M. Ehrmann, and W. Boos (1983) Periplasmic glycerophosphodiester phosphodiesterase of Escherichis coli, a new enzyme of the glp regulon. J. Biol. Chem. 258: 5428–5432.
Ignatova, Z., C. Hornle, A. Nurk, and V. Kasche (2002) Unusual signal peptide directs penicillin amidase from Escherichia coli to the tat translocation machinery. Biochem. Biophy. Res. Comm. 291: 146–149.
Widdick, D. A., K. Dilks, G. Chandra, A. Bottrill, M. Naldrett, M. Pohlschröder, and T. Palmer (2006) The twin-arginine translocation pathway is a major route of protein export in Streptomyces coelicolor. Proc. Natl. Acad. Sci. 103: 17927–17932.
Kieran, D., R. W. Rose, E. Hartmann, and M. Pohlschröder (2003) Prokaryotic utilization of the twin-arginine translocation pathway: a genomic survey. J. Bacteriol. 185: 1478–1483.
Chi, W. J., J. H. Song, E. A. Oh, S. W. Park, Y. K. Chang, E. S. Kim, and S. K. Hong (2009) Medium optimization and application of affinity column chromatography for trypsin production. J. Microbiol. Biotechnol. 19: 1191–1196.
Gauthier, C., H. Li, and R. Morosoli (2005) Increase in xylanase production by Streptomyces lividans through simultaneous use of the Sec- and Tat-dependent protein export systems. Appl. Envrion. Microbiol. 71: 3085–3092.
Kim, M. R., Y. H. Choeng, W. J. Chi, D. K. Kang, and S. K. Hong (2010) Heterologous production of streptokinase in secretory form in Streptomyces lividans and in nonsecretory form in Escherichia coli. J. Microbiol. Biotechnol. 20: 132–137.
Blaudeck, N., P. Kreutzenbeck, M. Müller, G. A. Sprenger, and R. Freudl (2005) Isolation and characterization of bifunctional Escherichia coli TatA mutant proteins that allow efficient tatdependent protein translocation in the absence of TatB. J. Biol. Chem. 280: 3426–3432.
Alami, M., I. Lüke, S. Deitermann, G. Eisner, H. G. Koch, J. Brunner, and M. Müller (2003) Differential interactions between a twin-arginine signal peptide and its translocase in Escherichia coli. Mol. Cell. 12: 937–946.
De Keersmaeker, S., L. Van Mellaert, E. Lammertyn, K. Vrancken, J. Anne, and N. Geukens (2005) Functional analysis of TatA and TatB in Streptomyces lividans. Biochem. Biophys. Res. Commun. 335: 973–982.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Chi, WJ., Oh, E.A., Kim, JH. et al. Enhancement of protein secretion by TatAC overexpression in Streptomyces griseus . Biotechnol Bioproc E 16, 59–71 (2011). https://doi.org/10.1007/s12257-010-0382-7
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s12257-010-0382-7