Production of <Emphasis Type="Italic">Chryseobacterium proteolyticum</Emphasis> protein-glutaminase using the twin-arginine translocation pathway in <Emphasis Type="Italic">Corynebacterium glutamicum</Emphasis> |
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Authors: | Yoshimi Kikuchi Hiroshi Itaya Masayo Date Kazuhiko Matsui Long-Fei Wu |
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Institution: | (1) Institute of Life Sciences, Ajinomoto Co., Inc., 1–1 Suzuki-cho, Kawasaki-ku, Kawasaki 210-8681, Japan;(2) Laboratoire de Chimie Bactérienne, UPR9043, Institut de Biologie Structurale et Microbiologie, CNRS, 13402 Marseille, France |
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Abstract: | The protein glutaminase (PG) secreted by the Gram-negative bacterium Chryseobacterium proteolyticum can deamidate glutaminyl residues in several substrate proteins, including insoluble wheat glutens. This enzyme therefore
has potential application in the food industry. We assessed the possibility to produce PG containing a pro-domain in Corynebacterium glutamicum which we have successfully used for production of several kinds of proteins at industrial-scale. When it was targeted to
the general protein secretion pathway (Sec) via its own signal sequence, the protein glutaminase was not secreted in this
strain. In contrast, we showed that pro-PG could be efficiently produced using the recently discovered twin-arginine translocation
(Tat) pathway when the typical Sec-dependent signal peptide was replaced by a Tat-dependent signal sequence from various bacteria.
The accumulation of pro-PG in C. glutamicum ATCC13869 reached 183 mg/l, and the pro-PG was converted to an active form as the native one by SAM-P45, a subtilisin-like
serine protease derived from Streptomyces albogriseolus. The successful secretion of PG via this approach confirms that the Tat pathway of C. glutamicum is an efficient alternative for the industrial-scale production of proteins that are not efficiently secreted by other systems. |
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Keywords: | Corynebacterium glutamicum Protein secretion Twin-arginine Translocation pathway Protein glutaminase |
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