Difference between revisions of "Part:BBa K2933155"
 
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'''Figure 1.''' Left: The result of PCR, Right:The result of double enzyme digestion verification.LaneM,Marker, Lane1, the plasmid with VIM-66, Lane2, after double enzyme verification
 
'''Figure 1.''' Left: The result of PCR, Right:The result of double enzyme digestion verification.LaneM,Marker, Lane1, the plasmid with VIM-66, Lane2, after double enzyme verification
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===References===
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[1]Yoshihiro Yamaguchi. Wanchun Jin. Kazuyo Matsunaga. Crystallographic investigation of the inhibition mode of a VIM-2 metallo-beta-lactamase from Pseudomonas aeruginosa by a mercaptocarboxylate inhibitor. J. Med. Chem.200750266647-6653
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[2]Biochemical, Mechanistic, and Spectroscopic Characterizationof Metallo-β-lactamase VIM‑2[J]. Biochemistry, 2014, 53(46):7321-7331.
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[3]Christopeit T , Carlsen T J , Helland R , et al. Discovery of novel inhibitor scaffolds against the metallo-β-lactamase VIM-2 by SPR based fragment screening[J]. Journal of Medicinal Chemistry, 2015:151017114758002.
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[4]Christopeit T , Yang K W , Yang S K , et al. The structure of the metallo-β-lactamase VIM-2 in complex with a triazolylthioacetamide inhibitor[J]. 2016.

Latest revision as of 11:13, 24 September 2019


His+Linker f+VIM-66

This part encodes the fusion protein of His tag and VIM-66 to promote the expression and purification of target protein(VIM-66).

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 51
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 824
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Usage and Biology

This composite part is made up with three basic parts, the HiS tag, the thrombin restriction site and our target protein VIM-66. It encodes a protein which is VIM-66 fused with His tag. The fusion protein is about 28.3 kD. It is convenient for us to purify our target protein.

Molecular cloning

We insert VIM-66 gene into the standard vector then transfer it into E.coli.

VIM-66-PCR.jpeg

Figure 1. Left: The result of PCR, Right:The result of double enzyme digestion verification.LaneM,Marker, Lane1, the plasmid with VIM-66, Lane2, after double enzyme verification

References

[1]Yoshihiro Yamaguchi. Wanchun Jin. Kazuyo Matsunaga. Crystallographic investigation of the inhibition mode of a VIM-2 metallo-beta-lactamase from Pseudomonas aeruginosa by a mercaptocarboxylate inhibitor. J. Med. Chem.200750266647-6653

[2]Biochemical, Mechanistic, and Spectroscopic Characterizationof Metallo-β-lactamase VIM‑2[J]. Biochemistry, 2014, 53(46):7321-7331.

[3]Christopeit T , Carlsen T J , Helland R , et al. Discovery of novel inhibitor scaffolds against the metallo-β-lactamase VIM-2 by SPR based fragment screening[J]. Journal of Medicinal Chemistry, 2015:151017114758002.

[4]Christopeit T , Yang K W , Yang S K , et al. The structure of the metallo-β-lactamase VIM-2 in complex with a triazolylthioacetamide inhibitor[J]. 2016.