Difference between revisions of "Part:BBa K2235007"

(→‎Important parameters)
(→‎Important parameters)
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|Optimal temperature
 
|Optimal temperature
 
|37 °C
 
|37 °C
−
|-
 
−
|rowspan="6"|Purification
 
 
|-
 
|-
 
|Number of amino acids
 
|Number of amino acids
−
|678
+
|XXX
 
|-
 
|-
 
|Molecular weight
 
|Molecular weight
−
|74.59 kDa
+
|XXX;kDa
 
|-
 
|-
−
|Theoretical pI
+
|rowspan="6"|Purification
−
|5.59
+
 
|-
 
|-
 
|Extinction coefficient at 280 nm <br> measured in water [M<sup>-1</sup> cm<sup>-1</sup>]
 
|Extinction coefficient at 280 nm <br> measured in water [M<sup>-1</sup> cm<sup>-1</sup>]

Revision as of 13:06, 18 October 2017


OmpR promoter regulating the expression of sialidase enzyme

OmpR is an osmotic pressure promoter. It can sense changes in for example sucrose concentration. A higher concentration of sucrose will lead to a higher expression of the gene after the OmpR promoter. In this biobrick the gene after OmpR is an enzyme called sialidase. This enzyme cuts sialic acid (that's for example found in mucus). The sialidase produced by this biobrick has a HIS-tag on it to make it easy to purify.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal XhoI site found at 189
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 636
    Illegal NgoMIV site found at 711
    Illegal NgoMIV site found at 801
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1181


Usage and biology

OmpR is an already well characterized biobrick (BBa_R0082). It is a promoter that takes advantage of the OmpR/EnvZ two-component system, which is utilized by many bacteria, to sense the change in osmotic pressure. We connected this promoter to the coding sequence for the enzyme sialidase. This enzyme hydrolyzes glycosidic linkages of terminal sialic acid residues in glycoproteins. This composite biobrick can be used to produce sialidase when the osmotic pressure in the environment around the bacteria increases.

Important parameters

Experiment Characteristic Result
Expression
Compatibility E. coli TOP10

E. coli ΔEnvZ

Promoter OmpR
Optimal temperature 37 °C
Number of amino acids XXX
Molecular weight XXX;kDa
Purification
Extinction coefficient at 280 nm
measured in water [M-1 cm-1] 		37400 (assuming all pairs of Cys residues form cystines)

36900 (assuming all Cys residues are reduced)

Tag C-terminal 6xHis
NAD+ detection
Limit of detection 2 nM