Difference between revisions of "Part:BBa K627011"

Revision as of 20:43, 21 September 2011

Fusion part of pBAD arabinose-inducible induction system and the HRV 14_3C protease
This BioBrick is a 2 part fusion of arabinose-inducible induction system and the HRV 14_3C protease. The recombinant type 14_3C protease from human rhinovirus (HRV 3C) recognizes the same cleavage site as the native enzyme: LeuGluValLeuPheGln↓GlyPro.
The small, 22 kDa size of the protease got its optimal activity at 4°C but is still very active at 37°C. It is commonly used for an easy tag removal after the purification of recombinant proteins carrying his-tag. The 14_3C works with a catalytic triade, containing the amino acid residues Ser-Asp-His at its active site.

The arabinose-inducible induction system was amplified via PCR from the pBAD_iGEMexpress vector and fused via NgoMIV with the HRV 14_3C protease. An effective induction can be accomplished with 2 mM up to 25 mM arabinose.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
INCOMPATIBLE WITH RFC[21]
Illegal BglII site found at 1523
Illegal BglII site found at 1711
Illegal BamHI site found at 1144
23
COMPATIBLE WITH RFC[23]
25
INCOMPATIBLE WITH RFC[25]
Illegal NgoMIV site found at 1239
1000
INCOMPATIBLE WITH RFC[1000]
Illegal SapI site found at 961

@@ Line 1: / Line 1: @@
 __NOTOC__
 <partinfo>BBa_K627011 short</partinfo><br>
-This construct contains the cleavage site for the 14_3C protease flanked by the sequence of beta-lactamase and the TorA signal sequence for the TAT (twin arginine transporter) pathway, thus in case of the expression of the complete construct the beta lactamase can be transported into periplasm and provide ampicillin resistance. The TorA signal-sequence originate from the enzyme Trimethylamin-N-Oxid-Reduktase , which needs to be folded in the cytoplasm. The beta lactamase is an enzyme which cleaves lactam rings and makes bacteria resistant to antibiotics like ampicillin and penicillin. The cleavage site for the HRV 14 3C protease was created via oligo hybridization and ligated into the construct with cleavage site for XhoI and NheI, which makes this part modular and easy adaptable for any other protease.
+This BioBrick is a 2 part fusion of arabinose-inducible induction system and the HRV 14_3C protease. The recombinant type 14_3C protease from human rhinovirus (HRV 3C) recognizes the same cleavage site as the native enzyme: LeuGluValLeuPheGln↓GlyPro.<br>
+The small, 22 kDa size of the protease got its optimal activity at 4°C but is still very active at 37°C. It is commonly used for an easy tag removal after the purification of recombinant proteins carrying his-tag. The 14_3C works with a catalytic triade, containing the amino acid residues Ser-Asp-His at its active site.<br>
+<br>
+The arabinose-inducible induction system was amplified via PCR from the pBAD_iGEMexpress vector and fused via NgoMIV with the HRV 14_3C protease. An effective induction can be accomplished with 2 mM up to 25 mM arabinose.<br>
 <!-- Add more about the biology of this part here