Functional analysis of the twin-arginine translocation pathway in Corynebacterium glutamicum ATCC 13869.
Compared to those of other gram-positive bacteria, the genetic structure of the Corynebacterium glutamicum Tat system is unique in that it contains the tatE gene in addition to tatA, tatB, and tatC. The tatE homologue has been detected only in the genomes of gram-negative enterobacteria. To assess the function of the C. glutamicum Tat pathway, we cloned the tatA, tatB, tatC, and tatE genes from C. glutamicum ATCC 13869 and constructed mutants carrying deletions of each tat gene or of both the tatA and tatE genes. Using green fluorescent protein (GFP) fused with the twin-arginine signal peptide of the Escherichia coli TorA protein, we demonstrated that the minimal functional Tat system required TatA and TatC. TatA and TatE provide overlapping function. Unlike the TatB proteins from gram-negative bacteria, C. glutamicum TatB was dispensable for Tat function, although it was required for maximal efficiency of secretion. The signal peptide sequence of the isomaltodextranase (IMD) of Arthrobacter globiformis contains a twin-arginine motif. We showed that both IMD and GFP fused with the signal peptide of IMD were secreted via the C. glutamicum Tat pathway. These observations indicate that IMD is a bona fide Tat substrate and imply great potential of the C. glutamicum Tat system for industrial production of heterologous folded proteins.
Kikuchi
Yoshimi
Y
Institute of Life Sciences, Ajinomoto Co., Inc., 1-1 Suzuki-cho, Kawasaki-ku, Kawasaki 210-8681, Japan. yoshimi_kikuchi@ajinomoto.com
Date
Masayo
M
Itaya
Hiroshi
H
Matsui
Kazuhiko
K
Wu
Long-Fei
LF
D000595:Amino Acid Sequence; D001173:Arthrobacter / Q000201:enzymology / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D048230:Corynebacterium glutamicum / Q000235:genetics / Q000378:metabolism*; D003910:Dextranase / Q000378:metabolism; D029968:Escherichia coli Proteins; D017353:Gene Deletion; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007218:Industrial Microbiology / Q000379:methods; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D011134:Polysaccharides / Q000378:metabolism; D017510:Protein Folding; D017422:Sequence Analysis, DNA
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D011134:Polysaccharides; C408585:twin-arginine translocase complex, E coli; D049452:Green Fluorescent Proteins; C008315:maltodextrin; D003910:Dextranase