Difference between revisions of "Part:BBa K3051421"
 
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<partinfo>BBa_K3051421 short</partinfo>
 
<partinfo>BBa_K3051421 short</partinfo>
  
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This is a lipase found in a metagenomics search of a compost done by the University of Gottingen. It was found to break down the ester bond of p nitrophenol octanoate. It has a similarity (77% identity) to Thermostable Lipase A (BBa_K258006), and we believe it has the same function. Its conserved domains include a hydrolase site and an RTX toxin related Ca2+ binding site, which may make it sensitive to Ca ions or act as a cofactor.  
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<h2>Comparative Enzyme Activity Assay</h2>
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<div align="center"><img height="85%" width="85%" src="https://2019.igem.org/wiki/images/8/83/T--Warwick--2019-EnzymeLipaseGraph.png"></img></div>
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<p><b>Figure 1. Relative Comparaison Lipase Activity Assay.</b> Candida Antarctica Lipase A, Bacillus Subtilis Lipase and Compost Lipase enzyme activity were examined using p-Nitrophenol octanoate as a substrate.</p>
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Figure 1 compares enzyme activity of Candida Antarctica Lipase A <bbpart>BBa_K3051001</bbpart>, Bacillus Subtilis Lipase <bbpart>BBa_K3051005</bbpart> and Compost Lipase <bbpart>BBa_K3051421</bbpart>. The lipase parts were tested and characterized using a [https://2019.igem.org/Team:Warwick/Results p nitrophenol octanoate assay]. The enzyme activity was determined by measuring change in light absorbance at a wavelength of 400nm, which directly correlated to the concentration of p-Nitrophenol (pNP), caused by lipolysis of p-Nitrophenol octanoate (pNPO) substrate. The data from the pNPO lipolysis assay defined the enzyme activity.
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The lipase was found in a metagenomics search of a compost performed by the University of Gottingen. The enzyme showed 77% identity to Thermostable Lipase A <bbpart>BBa_K258006</bbpart>, and likely conserves a similar function. The conserved domains include a hydrolase site and an RTX toxin related Ca2+ binding site - which could either cause sensitivity to Calcium ions or Calcium ions could act as a cofactor.  
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<div align="center"><img height="50%" width="50%" src="https://2019.igem.org/wiki/images/a/a1/T--Warwick--CMLP_3D_structure.png"></img></div>
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<p><b>Figure 2. CLMP 3D Structure.</b> The protein has a kDa of 64.6 and an estimated molar extinction coefficient of 75290 M-1cm-1 (calculated using ProtoPram ExPaSy tool); the following 3D structure (calculated using Phyre 2).</p>
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It has a kDa of 64.6 and an estimated molar extinction coefficient of 75290 M-1cm-1 (calculated using ProtoPram ExPaSy tool). It has the following 3D structure (calculated using Phyre 2)
 
  
 
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Latest revision as of 03:52, 22 October 2019


Compost Lipase

Comparative Enzyme Activity Assay

Figure 1. Relative Comparaison Lipase Activity Assay. Candida Antarctica Lipase A, Bacillus Subtilis Lipase and Compost Lipase enzyme activity were examined using p-Nitrophenol octanoate as a substrate.

Figure 1 compares enzyme activity of Candida Antarctica Lipase A BBa_K3051001, Bacillus Subtilis Lipase BBa_K3051005 and Compost Lipase BBa_K3051421. The lipase parts were tested and characterized using a p nitrophenol octanoate assay. The enzyme activity was determined by measuring change in light absorbance at a wavelength of 400nm, which directly correlated to the concentration of p-Nitrophenol (pNP), caused by lipolysis of p-Nitrophenol octanoate (pNPO) substrate. The data from the pNPO lipolysis assay defined the enzyme activity.

The lipase was found in a metagenomics search of a compost performed by the University of Gottingen. The enzyme showed 77% identity to Thermostable Lipase A BBa_K258006, and likely conserves a similar function. The conserved domains include a hydrolase site and an RTX toxin related Ca2+ binding site - which could either cause sensitivity to Calcium ions or Calcium ions could act as a cofactor.


Figure 2. CLMP 3D Structure. The protein has a kDa of 64.6 and an estimated molar extinction coefficient of 75290 M-1cm-1 (calculated using ProtoPram ExPaSy tool); the following 3D structure (calculated using Phyre 2).


Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Unknown
  • 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]