Difference between revisions of "Part:BBa K3007000"

Revision as of 16:35, 19 October 2019

HucR binding site

It is a special DNA sequence which can interact with HucR protein and repress the expression of downstream gene when uric acid is absent. If uric acid presents, the formation of HucR-hucO complex can be reduced and restore downstream gene expression. In our design, we used the HucR-hucO system as a detector for uric acid.

Contribution: QHFZ-China 2019

Group: QHFZ-China
Authors: Cheng Li
Introduction:

   HucR is a novel uric acid-responsive member of the MarR Family from Deinococcus radiodurans [1]. When uric acid is absent, HucR can bind as a homodimer to a single operator site on DNA (hucO) with very high affinity (Kd = 0.29 ± 0.02 nM). We located this part in a modified lac promoter, which contains a HucR binding site (hucO) and a mutate lacO sequence (BBa_K3007001). Therefore, this promoter could be under the control of HucR, instead of LacI. If uric acid presents in high concentration, HucR will release from hucO and the expression of downstream gene will recover from the inhibition. In our design, HucR-hucO system works as a uric acid concentration detector.

Design:
Figure 1. Schematic cartoon of the DNA construct of BBa_K3007000
Documentation:

   This year, QHFZ-China designed a UA monitor system in E. coli (Fig. 1). Pc is a constitutive promoter, Pcp6 promoter, and it promotes the expression of HucR and YgfU. If the concentration of uric acid (UA) in environment is low, HucR will bind to hucO sequence (located in PhucR), which inhibits the expression of downstream reporter, dsRed or sfGFP. When extracellular UA is present, YgfU can transport UA into the cytoplasm, which leads HucR dissociates from hucO, and induces the fluorescent protein expression.

File:T--QHFZ-China--BBa K3007000 2

Figure 1. Working mechanism of the uric acid detection system in E. coli. (A) Schematic diagram of the gene circuit design. (B) Structure of PhucR, showing the location of BBa_K3007000.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

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      This year, QHFZ-China designed a UA monitor system in E. coli (Fig. 1). Pc is a constitutive promoter, Pcp6 promoter, and it promotes the expression of HucR and YgfU. If the concentration of uric acid (UA) in environment is low, HucR will bind to hucO sequence (located in PhucR), which inhibits the expression of downstream reporter, dsRed or sfGFP. When extracellular UA is present, YgfU can transport UA into the cytoplasm, which leads HucR dissociates from hucO, and induces the fluorescent protein expression.
-[[File:T--Valencia_UPV--sfGFP_spectrum.png|300px|thumb|none|alt=sfGFP spectra.|Figure 1. Working mechanism of the uric acid detection system in E. coli. (A) Schematic diagram of the gene circuit design. (B) Structure of PhucR, showing the location of BBa_K3007000.]]
+[[File:T--QHFZ-China--BBa_K3007000_2|450px|thumb|none|Figure 1. Working mechanism of the uric acid detection system in E. coli. (A) Schematic diagram of the gene circuit design. (B) Structure of PhucR, showing the location of BBa_K3007000.]]