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| Prokaryotic Expression of CM of T-protein from E.coli and Its Crystal Optimization Guided by NMR |
| RUI Bin, XIE Chang-lin, JIANG Na, WEN Han |
| School of Life Sciences, Anhui Agricultural University, Hefei 230036, China |
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Abstract More and more researches have shown that the pathogenicities of many bacteria are related to their chorismate mutases in the shikimate pathway. CMs represent a hot selective target for the development of antibiotics and fungicides. By PCR the recombinant CMT was expressed in competent cells E.coli BL21-Gold(DE3) successfully. The purity could reach electrophoresis purity after purification by Ni-NTA affinity chromatography, GE superdex75 gel filtration column and GE MonoQ cation-exchange chromatography. The result of mass spectra matched with the original sequence of CMT basically. And then CMT crystal were found in the condition of 30% PEG 4000,100mmol/L NaAc pH4.6,200mmol/L NH4Ac. The application of 1H-15N HSQC NMR provided guidance for optimization of CMT crystal, and finally we successfully obtained the high diffraction CMT crystal. This work lays the foundation for the structure of CMT.
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Received: 17 April 2013
Published: 25 September 2013
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