Team:Hong Kong CUHK/backgroundPAH

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<div id="partable">
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     <h1>Background</h1><p3>
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     <p2>
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<h3>Background</h3>
     <p>Our project initiated from dealing with toxic chemicals found in second-hand smoke (or environmental  tobacco smoke, ETS) and cooking fume. After carefully examining the composition  of pollutes, we noticed that there was one group of highly toxic and  carcinogenic compounds, polycyclic aromatic hydrocarbons (PAHs). The toxicity of PAHs is difficult  to handle, and many scientists are doing intensive research on this issue.</p>
     <p>Our project initiated from dealing with toxic chemicals found in second-hand smoke (or environmental  tobacco smoke, ETS) and cooking fume. After carefully examining the composition  of pollutes, we noticed that there was one group of highly toxic and  carcinogenic compounds, polycyclic aromatic hydrocarbons (PAHs). The toxicity of PAHs is difficult  to handle, and many scientists are doing intensive research on this issue.</p>
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    <p>PAHs are  notorious for its harm to both environment and human health. PAHs commonly  appear in people&rsquo;s daily life, from second-hand smoke  to cooking fume.  Also, PAHs can be generated in a large amount by  incomplete combustion of carbon-containing fuels. (Li et al. 2003, Zhang et al.  2013) PAHs may dissolve in air, diffuse in water, or precipitate in soil,  causing large-scale,  wide-spread pollution. (Samanta, Singh, and Jain 2002).Furthermore, most of the PAHs is recorded to be carcinogenic, mutagenic, or  teratogenic. (Li et al. 2003, Gauggel-Lewandowski et al. 2013) For example, the  first and foremost study of carcinogenicity of benzo[a]pyrene (BAP), a compound belongs to PAHs  family, dates back to the observation that chimney  sweepers developed scrotum cancer easier than others in 18th century.  (Boffetta, Jourenkova, and Gustavsson 1997)</p>
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<p>PAHs are  notorious for its harm to both environment and human health. PAHs commonly  appear in people&rsquo;s daily life, from second-hand smoke  to cooking fume.  Also, PAHs can be generated in a large amount by  incomplete combustion of carbon-containing fuels. (Li et al. 2003, Zhang et al.  2013) PAHs may dissolve in air, diffuse in water, or precipitate in soil,  causing large-scale,  wide-spread pollution. (Samanta, Singh, and Jain 2002).Furthermore, most of the PAHs is recorded to be carcinogenic, mutagenic, or  teratogenic. (Li et al. 2003, Gauggel-Lewandowski et al. 2013) For example, the  first and foremost study of carcinogenicity of benzo[a]pyrene (BAP), a compound belongs to PAHs  family, dates back to the observation that chimney  sweepers developed scrotum cancer easier than others in 18th century.  (Boffetta, Jourenkova, and Gustavsson 1997)</p>
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     <p>With the  power of synthetic biology, we aimed to develop a  method using E. coli to degrade PAHs into other non-toxic chemicals.</p></p3>
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     <p>With the  power of synthetic biology, we aimed to develop a  method using E. coli to degrade PAHs into other non-toxic chemicals.</p>
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    <h3>Degradation Pathway</h3>
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    <p>To degrade PAHs, we used a combination of two different codon-optimized  enzymes, namely laccase from <em>Bacillus sp.</em> HR03 and catechol 1,2-dioxygenase from <em>Pseudomonas  putid</em>a KT2440, in <em>Escherichia coli</em>.  After steps such as oxidation and ring-cleavage, PAHs can be degraded into more  simple and less toxic chemicals (figure: proposed degradation pathway).</p>
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    <p>&nbsp;</p>
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<p align="center"><img src="https://static.igem.org/mediawiki/2013/0/05/Pah5.png" alt="5" width="591" height="194"></p>
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    <p><strong>Laccase</strong><br>
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Laccase involved in the first step of degradation pathway. Hadibarata <em>et al</em>.[2] proposed that laccase could  play a significant role in benzo[a]pyrene (BaP) degradation. It was suggested  that laccase would add oxygen atoms onto the ring, changing PAHs into  quinone-like intermediates, which are easier for degradation, due to their  higher water solubility and weaker phenolic structures [2, 9]. Among various  bacterial laccases, we chose the laccase from <em>Bacillus sp</em>. HR03. The optimal conditions for this laccase are pH7  and 70 C, with acceptable activity in lower temperature such as 20 ºC [6].  Therefore, it is suitable for our experiment.</p>
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    <p><strong>Catechol 1,2-dioxygenase</strong><br>
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Followed by the action of laccase, catechol 1,2-dioxygenase from <em>Pseudomonas putida</em> KT240 continues  further degradation on the quinone-like compound [5]. Dioxygenase is an  oxidative enzyme which induces ortho-ring cleavage of catechol, a phenolic intermediate  of several metabolisms related to aromatic compound degradations [1, 5].  Moreover, a previous study indicated that 1,2-dioxygenase may play an important  role in BaP degradation [2]. Therefore, this enzyme was included for the  proposed metabolic pathway of PAHs in the project. The optimal conditions for  catechol 1,2-dioxygenase are pH7.5-8 and 25-30 ºC [5].</p>
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    <p><strong>QsrR</strong><br>
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To regulate PAHs  degradation, we used quinone sensing and response repressor (QsrR).&nbsp;QsrR  is a transcriptional regulator under the thiol-stress-sensing regulator YodB  family in <em>Staphylococcus aureus</em> [4].  In our proposed pathway, the intermediates, quinone-like compounds, can  function as the signaling molecules. Once bind with quinone molecules, QsrR  would leave the target DNA, and thus allow&nbsp;the expression of downstream  enzymes for subsequent degradations. With the regulation from QsrR, <em>E.coli</em> can avoid producing unnecessary  enzymes under PAHs-free situation, and therefore make them more viable.
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    </p3>
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Latest revision as of 16:01, 28 October 2013

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