• BMB Reports
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• v.38 no.5
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• pp.624-631
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• 2005

Tuberculosis, caused by Mycobacterium tuberculosis, continues to be one of the main diseases to mankind. It is urgent to discover novel drug targets for appropriate antimicrobial agents against this human pathogen. The shikimate pathway is onsidered as an attractive target for the discovery of novel antibiotics for its essentiality in bacteria and absence in mammalian cells. The Mycobacterium tuberculosis aroE-encoded shikimate dehydrogenase was cloned, expressed and purified. Sequence alignment analysis shows that shikimate dehydrogenase of Mycobacterium tuberculosis exhibit the pattern of G-X-(N/S)-V-(T/S)-X-PX-K, which is highly conserved within the shikimate dehydrogenase family. The recombinant shikimate dehydrogenase spectrum determined by CD spectroscopy showed that the percentages for $\alpha$-helix, $\beta$-sheet, $\beta$-turn, and random coil were 29.2%, 9.3%, 32.7%, and 28.8%, respectively. The enzymatic characterization demonstrates that it appears to be fully active at pH from 9.0 to 12, and temperature $63^{\circ}C$. The apparent Michaelis constant for shikimic acid and $NADP^+$ were calculated to be about $29.5\;{\mu}M$ and $63\;{\mu}M$. The recombinant shikimate dehydrogenase catalyzes the substrate in the presence of $NADP^+$ with an enzyme turnover number of $399\;s^{-1}$. Zymological studies suggest that the cloned shikimate dehydrogenase from M. tuberculosis has a pretty activity, and the work should help in the discovery of enzyme inhibitors and further of possible antimicrobial agents against Mycobacterium tuberculosis.

• Korean Journal of Weed Science
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• v.15 no.2
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• pp.148-153
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• 1995

Glyphosate (N-[phosphonomethyl]glycine) applied to the assimilate-exporting leaves or sprayed to the whole plants of tomato(Lycopersicon esculentum Mil var. Moneymaker) induced the rapid inhibition of 5-enolpyruvyl skimic acid 3-phosphate synthase(EPSP-synthase). It shows that EPSP-synthase activity precedes chlorophyll loss. There is no difference in EPSP-synthase activity between in vivo tomato meristem and cell suspension culture if glyphosate is not applied. The EPSP-synthase activity is in a range of 4 to 6 nkat per mg protein. The inhibition of EPSP-synthase action is induced within 36 h after glyphosate application while the Chl contents were reduced 48 h after the application. In cell suspension culture of tomato and Corydalis (Corydalis sempervirens), a sublethal concentration of glyphosate retards the fresh weight increase and prolonged lag phase. The fresh weight is reached maximal about 14 days after the subculture in the presence of glyphosate. The inhibitory effect of glyphosate on EPSP-synthase is remarkably induced in lag phase.

• Journal of Plant Biotechnology
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• v.34 no.3
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• pp.253-261
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• 2007

This study was conducted to develop an antibiotics marker-free potato (Solanum tuberosum L., cv. Taedong valley) plant having resistance against two herbicides. Agrobacterium tumefaciens strain EHA105, harboring a binary vector plasmid pCAMBIA3300 containing bar gene under the control of a promoter CaMV35S and linked CP4-EPSPS genes driven by CaMV35S promoter, was used in the current study. The leaf segments of newly bred potato variety (cv. Taedong Valley) was co-cultured with Agrobacterium. Then, the regenerated individual shoots were excised and transferred to potato multiplication medium supplemented with 0.5 mg/L phosphinothricin. The shoots were rooted in MS medium without hormone and obtained putative transgenic plant E3-6. Integration of target genes into the E3-6 plant and their expression was confirmed by PCR, Southern analysis, and ELISA test. The tissue necrosis test on young leaf blade and shikimic acid accumulation test using the tissue of E3-6 plant were conducted to investigate the resistance to glufosinate-ammonium and glyphosate, respectively. The transgenic plants (E3-6) simultaneously showed a high resistance to both herbicides. The same results were surely obtained also in the whole plants foliar-treated with alone or mixture of two herbicides, glufosinate-ammonium and glyphosate.