Team:IIT Delhi/project
Research
Since our target was to reduce air pollution, we targeted genes that act on NOx and SOx gases. Research was done extensively and the following genes were found to be best fitting for our project-
Types of Heme : The most common types of heme are heme b and heme c. Heme b: It is iron-protoporphyrin IX that is bound noncovalently to protein Heme c: heme c has two covalent thioether bonds formed between Cys side chains and the heme vinyl groups at positions 2 and 4 . The stereochemistry of heme attachment is the same in all known examples of heme c, and the vinyl groups at positions 2 and 4 are attached to the N- and C-terminal Cys of the CXXCH motif, respectively.
Other, less common, derivatives of heme include heme d1, present in cytochrome cd1 nitrite reductase, heme a, found in cytochrome c oxidase, and the related heme o, found in some bacterial oxidases. From a chemical perspective, hemes b and c are very similar to each other and thus are not expected to display significant inherent differences in electronic structure or reduction potential. From a biosynthetic perspective, however, these types of heme are quite different; because heme c is synthesized from heme b, the use of heme c requires a greater investment from the organism. The heme which is found in NrfA is Heme c Heme Biosynthesis Pathway :
Cytochrome c biogenesis: Natural modifications to heme occur after the ferrous iron (Fe2) is inserted into the porphyrin by ferrochelatase, the last enzyme in heme biosynthesis CCm complex : It is a complex of Transmembrane protein which helps in Heme maturation .From modelling we believe that by upregulating CCm complex of proteins we can increase heme incorporation into our proteins.
Technique to improve heme incorporation: V - Sulphite Reductase(Sir)- NADPH-sulfite reductase (SiR) of Escherichia coli catalyzes the reduction of sulfite to sulfide and is required for synthesis of L-cysteine from inorganic sulfate The native enzyme has a subunit structure a8B4 , where a8 is a flavoprotein (SiR-FP) containing both flavin adenine dinucleotide and flavin mononucleotide and P is a hemoprotein (SiR-HP) containing an Fe4S4 centre and a single molecule of siroheme Electron flow between these cofactors proceeds from NADPH to flavin adenine dinucleotide to flavin mononucleotide in the flavoprotein, then to a closely coupled Fe4S4 siroheme center in the hemoprotein, and finally from siroheme to sulphiteThe SiR-FP and SiR-HP components of SiR are encoded by cysJ and cysl, respectively. These genes are contiguous and together with cysH, the gene for 3'-phosphoadenosine 5'-phosphosulfate sulfotransferase, comprise an operon with the gene order promoter-cysJ-cysI-cysH cysJIH operon is part of the positively regulated cysteine regulon (15) and requires sulfur limitation CysB protein, and either O-acetyl-L-serine or N-acetyl-L-serine for expression SiR activity is also dependent on cysG, which encodes a uroporphyrinogen III methyltransferase necessary for the synthesis of siroheme This gene is located more than 10 min away from cysJIH on the chromosomal map (34) and is not tightly regulated as part of the cysteine regulatory VI - References- |
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Prototype
Why the "Pollution Crusader Device"? RSPM and NOx when combined, form a deadly combination which drastically affects human health. RSPM when inhaled acts like an abrasive and wears the lining of respiratory tract exposing soft spots where NOx reacts and causes severe issues. So we decided to eliminate soot and NOx as the primary objective. Analysis of soot: Soot is of two types: hydrophilic as well as hydrophobic. Hydrophilic soot dissolves in water, whereas hydrophobic soot dissolves in organic solvents like alcohol. To get rid of both the hydrophilic and hydrophobic components, we decided to use a solution of acetic acid in water as the solvent. Acetic acid, being an organic solvent, dissolves the hydrophobic part, and water, dissolves the hydrophobic soot. Using acetic acid, however, offers an added advantage as well. Being an acid, it reduces the PH of the medium and hence decreases the solubility of acidic NOx and SOx, from the exhaust into this solvent. Thus, the tank containing this solvent dissolves soot and NOx, Sox gases move out with the exhaust. The exhaust out of this tank is now passed into the desooter tank. The desooter tank has the following main design components:
A provision of tar exit is needed to remove this tar slurry continuously out of the tank. Spargers are needed to form bubbles of exhaust gas in the mixture. Blowers are also attached to the system to compensate for the back pressure generated in the system at each step. Now, we needed to condense the water vapor which flow along with exhaust gases out of this tank. This was required so that these excess water vapor do not clog the silica gel at the outlet of the tank. For this, we attach a heat exchanger at the outlet of the tank. It’s a cross flow type heat exchanger, with which we did some slight innovations to increase the efficiency of the condensation process. Cold water flows through the exchanger pipes and steam flows on the outside. We here filled the outside of the pipe portion of the exchanger with metal ball bearings. These ball bearings, through conduction, cool down to acquire the temperature(similar to) of the flow pipes and hence increase the surface area available to the vapor for condensation, speeding up the condensation process. The remaining exhaust, congaing NOx, Sox and some amount of water vapor now passes over the silica gel tank at the outlet of this desooter tank. Studies showed that NOx and SOx, in the presence of water vapor, react over Alumina-silica gel(SiO2.Al203) to lead to an oxidation reduction reaction forming NO and H2SO4. This was a remarkable discovery as the NRFA bacteria we are working with, effectively reduce NO into harmless forms, and hence we needed NO as the output from this tank. Thus, the NO rich gas from this tank is passed into the next tank containing NRFA bacteria for further action. H2SO4 produced is extracted and used for industrial purposes. Click here to view the model FUTURE SCOPE OF THE PROJECT AND TARGETS TO BE ACHIEVED: To optimize the design of the desooter tank and the entire contraption, to increase efficiency and reduce the cost of the process. To ultimately make the process so cheap and efficient that it can replace SCR in big engines, industries and power plants. Use the soot (obtained as dissolved in water) to make ink to ultimately make the process self-sustainable and bring down the processing cost to negligible amounts by accounting for commercial selling of this ink obtained. This will also further reduce environmental repercussions as soot now is converted into a useful product, rather than being dumped into the environment To work on developing and optimizing the desooter tank design to reduce power consumption in the blower, as well as the heat exchanger. Hence, through optimization, we aim to finally reach a design which gets the engine to work efficiently at even higher trade-off points. Conventionally, diesel engines are operated at temperature ranges which are not very high, as operation at high temperature leads to higher amount of NOx in the exhaust. Thus, they are always operated at a trade-off temperature lower than these high temperature values, to reduce NOx emission, however, also leading to a decrease in the engine’s efficiency, and increased amounts of CO and soot emission (due to more unburnt C content in the exhaust now). SC-R’s are not efficient enough, and thus, do not lead to a significant increase in the trade-off temperature. Thus, if an efficient and cost effective design of this NRFA driven pollution crusader device is realized, it would bring about a remarkable change in the automobile and diesel engine industry, and increase the efficiency of these devices, while reducing both the cost, and carbon footprints on the environment. |
Conclusion
While working in the project, towards the end, the main complexity we faced was of Heme protein. It is so because most of the enzymes we were using were Cytochrome C Hemo proteins, which require a Heme group for thein 100% activity. While scavenging the research papers, we found out that nobody, till date had found a perfect solution for the over expression of Heme protein. Under the constraints of time, we managed to compile the literature, designed the perfect strategy and using Copasi (modelling tool), we found some novel pathways to regulate and manipulate the increase in Heme production and hence the activity of the Heme proteins. We did manage to extensively characterize the NrfA cloned gene, which served as the nitrate reductase. We found out that even without using any extracellular or intracellular Heme over production, our clones were highly efficient in reducing the nitrite and subsequently converting it into ammonia. Finally, we came to the conclusion that while expressing NrfA in a constitutive fashion, there was increased conversion of NOx to ammonia. In the mechanical half of the project, we designed a prototype which in effect served as the perfect “Pollution Crusader”. We tested the model against the exhaust emitted by a diesel engine. Although the efficiency wasn’t that high, it did show positive results. We observed considerable reduction in the amount of NOx and SOx reduced. In a nutshell, our prototype model is capable of removing the harmful soot particles, gases like NOx and SOx and also the particulate matter present in the exhaust gases of the various engines being used. The prototype model designed will not only serve as an alternative for the existing technology, but has the potential to bring revolutionary changes in the upcoming days. |