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Contents |
1. Call for Biofuels, especially Alkanes
Nowadays, scientist have put great attention on biofuels, hoping to find a solution to energy crisis and climate change. Compared with fossil fuels, biofuels are renewable, and biofuel can be used indefinitely without any net carbon emissions [1]. Therefore, biofuels are promising candidates for mitigating dependence on diesel fuels.
Figure 1. Comparison of carbon cycles of fossil fuels and biofuels
Among all kinds of biofuels, alkanes stand out because of their excellent properties. First, alkanes have higher energy density, for example, enthalpy of combustion of pentadecane is approximately -47.0 MJ/kg compared with -29.7 MJ/kg for ethanol [2]. Then, compared with ethanols with a freezing point of -114℃,which alkanes have a higher freezing point of about -3~19℃, so they are more likely to be compatible with existing engines as well as transport and storage infrastructures. Besides, they can serve as drop-in replacement for fossil fuels.
2. Biosynthesis of alkanes
Pathways of long-chain alkane sythesis in microbes has been studied. Two enzymes, acyl-ACP reductase(AAR) and aldehyde decarbonylase(ADC), were heterologously expressed in E.coli to reduce fatty acyl-ACPs to corresponding aldehydes and then convert them to alkanes[3]. Fatty aldehydes can also be produced from fatty acids and fatty acyl-CoAs, catalyzed by carboxylic acid reductase(CAR) and acyl-CoA reductases(ACR) respectively, which has been identified in many species[4,5].
Figure 2. Typical alkane biosynthesis pathways
3. Sensing & Detecting alkanes
In the research about alkane biosynthesis, productivity of microbes and profile of alkanes produced are basic information that need to be acquired. In the process of directed evolution towards high producing bacteria, a selection strategy that relates alkane productivity with cell growth rate is crucial. In large-scale alkane production, it’s important to monitor the producing process at any time. To achieve all these goals, we need to detect and analyse alkanes in samples both qualitatively and quantitively.
However, current methods of alkane detection have many limits. First, extracting the product from original samples and operating high-tech equipments such as GC-MS makes the analysis process quite costly, time-consuming and laborious. Second, to select out target strains in directed evolution, commonly used screening tools are inherently low throughput. What's more, the current methods can hardly perform real-time in vivo detection for industrial production. We’re looking forward to developing an alkane sensor without these limits.
Figure 3. GC-MS, a high-tech equipment often used in alkane detection
To develop an alkane sensor, we need to find a mechanism which can respond to alkanes, our target products. In nature, many oil-degrading prokaryotes such as P. putida Gpo1, Alcanivorax borkumensis SK2 and Acinetobacter baylyi ADP1 have gene circuits responding to alkanes(alkS-alkB from P. putida Gpo1, alkS-alkB1 from Alcanivorax borkumensis SK2[6], and alkR-PalkM from Acinetobacter baylyi ADP1[2]). Among them, alkR-PalkM circuit in Acinetobacter baylyi ADP1 has remained untouched in iGEM competition. AlkR responds to a broad range of alkanes with carbon chain length from C7 to C36, and it’s the only bioreporter that is able to detect alkane with carbon chain length greater than C18[7]. Additionally, although side products such as fatty alcohol whose structure is similar to alkane can also combine with alkR, their combined compounds will inhibit PalkM, which makes this mechanism specifically suitable for alkane synthesis pathway.
References
[1] Yan Kung, Weerawat Runguphan, and Jay D. Keasling. “From Fields to Fuels: Recent Advances in the Microbial Production of Biofuels.” ACS Synth. Biol. 2012, 1, 498−513
[2] Mathew A Rude and Andreas Schirmer.“New microbial fuels: a biotech perspective.” Current Opinion in Microbiology 2009, 12:274–281
[3] Andreas Schirmer et al. “Microbial Biosynthesis of Alkanes.” Science 2010: Vol. 329 no. 5991 pp. 559-562
[4] Rebecca M Lennen and Brian F Pfleger “Microbial production of fatty acid-derived fuels and chemicals” Current Opinion in Biotechnology 2013, 21:1–10
[5] M. Kalim Akhtar, Nicholas J. Turner, and Patrik R. Jones. “Carboxylic acid reductase is a versatile enzyme for the conversion of fatty acids into fuels and chemical commodities.” PNAS January 2, 2013,vol. 110, no. 1, 87–92
[6] Rekha Kumari, Robin Tecon, Siham Beggah et al.(2011) “Development of bioreporter assays for the detection of bioavailability of long-chain alkanes based on the marine bacterium Alcanivorax borkumensis strain SK2.” Environmental Microbiology 13(10), 2808–2819
[7] Zhang, Dayi, et al.(2012) "Whole-cell bacterial bioreporter for actively searching and sensing of alkanes and oil spills." Microbial Biotechnology 5.1: 87-97
