期刊
APPLIED MICROBIOLOGY AND BIOTECHNOLOGY
卷 53, 期 3, 页码 282-288出版社
SPRINGER VERLAG
DOI: 10.1007/s002530050022
关键词
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The level of lysine-6-aminotransferase (encoded by the lat gene), an enzyme that commits lysine to the cephamycin biosynthesis pathway, is very low in wild type Nocardia lactamdurans . Two lat overexpression systems (pAMEXlat and pSAFlat) were constructed to express the promoterless fat gene of N. lactamdurans from the strong promoters amyP (of the alpha-amylase gene) and safP (of the secretion activating factor gene) of Streptomyces griseus. Both constructions led to very high levels of lysine-6-aminotransferase (between 8- and 15-fold) in the cells. Expression of (at from the amy) promoter was optimal in glycerol-containing medium and was negatively regulated by glucose. The high levels of lysine-6-aminotransferase resulted in a 50 200% increase in cephamycin C production in the standard fermentation conditions. Onset of cephamycin C biosynthesis occurred at the same time in control and in lat-overexpressing strains, but the cephamycin production rate was clearly higher in transformants overexpressing the lat gene. Furthermore, HPLC analysis of cephamycin C in the culture broths revealed an early depletion of biosynthetic intermediates and an accumulation of cephamycin C when the lat gene was overexpressed. These results indicate that lysine-6-aminotransferase activity is limiting for cephamycin C biosynthesis under some culture conditions.
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