Secretion of active xylanase C from Streptomyces lividans is exclusively mediated by the Tat protein export system.
The bacterial twin-arginine translocation (Tat) pathway transports folded proteins across the cytoplasmic membrane. The precursors targeted to the Tat pathway have signal peptides bearing the consensus motif (S/T-R-R-X-F-L-K). The xylanase C (XlnC) of Streptomyces lividans is a 20-kDa secreted enzyme. The XlnC signal peptide is 49 amino acids long and contains the S-R-R-G-F-L-G sequence, which is similar to the twin-arginine consensus motif. In S. lividans, XlnC secretion was impaired in a tatC insertion mutant, which is unable to secrete proteins that are dependent on the Tat system. When the signal peptide of XlnC was replaced by the Sec-dependent signal peptide of xylanase A, XlnC was secreted as an inactive form and demonstrated rapid proteolytic degradation in the culture supernatant, thus indicating that XlnC was specifically secreted through the Tat system. Deletions of the n-region of the XlnC signal sequence showed that a minimum of six amino acids residues preceding the twin-arginine motif was required to secrete XlnC. Replacement of one or both arginines by lysine residues in the twin arginine motif decreased four- and sevenfold, respectively, the enzyme production but did not abolish it. However, pulse chase experiments showed that the half-life of the precursor was from 2 to 3 h instead of 11 min for the wild- type precursor. Since XlnC is not associated with cofactors to exhibit activity, it is therefore a newly identified prokaryotic non-redox Tat substrate.
Faury
Damien
D
INRS-Institut Armand-Frappier, Université du Québec, 531 Blvd. des Prairies, Ville de Laval, Québec, Canada H7V 1B7.
Saidane
Saloua
S
Li
Haiming
H
Morosoli
Rolf
R
D002462:Cell Membrane; D016384:Consensus Sequence; D043364:Endo-1,4-beta Xylanases / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000378:metabolism*; D011498:Protein Precursors / Q000378:metabolism; D021382:Protein Sorting Signals; D021381:Protein Transport; D017384:Sequence Deletion; D013302:Streptomyces / Q000201:enzymology*
D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D011498:Protein Precursors; D021382:Protein Sorting Signals; C408585:twin-arginine translocase complex, E coli; D043364:Endo-1,4-beta Xylanases