title: Daunorubicin forms a specific complex with a secreted serine protease of Streptomyces peucetius.
abstract: Daunorubicin forms specific complex with an extracellular protease in the Streptomyces peucetius culture. The drug-protein complex co-migrates in non-denaturing PAGE as a red band. De novo peptide sequencing by nano-LC-ESI-MS/MS and MASCOT analysis identified the daunorubicin binding protein as serine protease precursor. The same protease precursor was purified sans the daunorubicin, from the mutant named ΔDPSAmut, which is deficient in daunorubicin production. Daunorubicin was added to ΔDPSAmut culture and the protease readily formed the daunorubicin-protease complex. Ability of serine protease precursor to form a selective complex with daunorubicin was confirmed by this study. Selective binding of protease to daunorubicin was seen as self-resistance determinant for the organism to survive toxic levels of the drug outside the cell. Daunorubicin-protease complex placed on S. peucetius lawn did not produce clearing zone around it, whereas daunorubicin purified from the complex did produce the clearing zone. Thereby it is concluded that the protease sequesters daunorubicin to prevent its entry into cells. Sequestration of daunorubicin by extracellular protease helps the organism to maintain a steady state sub-inhibitory level of drug around the cells. A new self-resistance determinant is reported here.
authors.affiliation: Department of Genetic Engineering, School of Biotechnology, Madurai Kamaraj University, Madurai, 625021, India.
authors.forename: Rashmi
authors.initials: R
authors.lastname: Dubey
authors.forename: Karuppasamy
authors.initials: K
authors.lastname: Kattusamy
authors.forename: Kuppamuthu
authors.initials: K
authors.lastname: Dharmalingam
authors.forename: Ranjan
authors.initials: R
authors.lastname: Prasad
mesh_terms: D003630:Daunorubicin / Q000378:metabolism*; D013058:Mass Spectrometry; D011485:Protein Binding; D057057:Serine Proteases / Q000378:metabolism*; D013302:Streptomyces / Q000201:enzymology* / Q000378:metabolism
chemical_list: D057057:Serine Proteases; D003630:Daunorubicin
references.citation: FEBS Lett. 1988 Mar 28;230(1-2):171-5
references.pmid: 2450783
references.citation: Can J Microbiol. 1985 Mar;31(3):287-94
references.pmid: 3859358
references.citation: Biotechnol Bioeng. 1969 Nov;11(6):1101-10
references.pmid: 5365804
references.citation: Nucleic Acids Res. 2003 Jul 1;31(13):3497-500
references.pmid: 12824352
references.citation: Appl Environ Microbiol. 2003 Jan;69(1):84-91
references.pmid: 12513980
references.citation: Mol Microbiol. 2005 Jan;55(2):624-36
references.pmid: 15659175
references.citation: FEMS Microbiol Lett. 2010 Apr;305(1):18-27
references.pmid: 20158521
references.citation: J Mol Biol. 1990 Oct 5;215(3):403-10
references.pmid: 2231712
references.citation: FEMS Microbiol Lett. 1998 Nov 15;168(2):243-9
references.pmid: 9835035
references.citation: Nucleic Acids Res. 1983 Nov 25;11(22):7911-25
references.pmid: 6316278
references.citation: Annu Rev Microbiol. 1989;43:207-33
references.pmid: 2679354
references.citation: J Bacteriol. 1997 Mar;179(5):1796-804
references.pmid: 9045843
references.citation: J Bacteriol. 1994 Oct;176(20):6270-80
references.pmid: 7928998
references.citation: Adv Appl Microbiol. 1987;32:203-14
references.pmid: 3307320
references.citation: Biochem Biophys Res Commun. 2003 Nov 14;311(2):460-4
references.pmid: 14592436
references.citation: Proc Natl Acad Sci U S A. 1991 Oct 1;88(19):8553-7
references.pmid: 1924314
references.citation: Gene. 1994 Dec 30;151(1-2):1-10
references.pmid: 7828855
references.citation: Eur J Biochem. 1999 Oct 1;265(1):308-17
references.pmid: 10491187