Part:BBa_M50444:Design
Modified FimH protein with affinity for heavy metals
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 320
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 541
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
It is a rhamnose inducible promotor, which when fully induced by 100mM seemed to have a high metabolic burden on our cell slightly inhibiting growth. Between our fimH sequence and our sfGFP and His-tag sequence there is a GGSG linker to provide space and try to minimize steric hinderance and misfolding of proteins. There is no linker between the His-tag and sfGFP, but we would recommend adding longer linkers and an additional linker between the sfGFP and His-tag. This was grown in DH5a because they are a type of E. coli that express the fim genes and the bp sequence was optimized for E. coli.
Source
BBa_M50080- strong RBS
BBa_K914003- promoter
BBa_K112703- His-tag
BBa_K1486053- GGSG linker
BBa_M50060- Terminator
BBa_I746916- sfGFP
References
https://parts.igem.org/Part:BBa_M50080
https://parts.igem.org/Part:BBa_K914003
https://parts.igem.org/Part:BBa_K112703
https://parts.igem.org/Part:BBa_K1486053
https://parts.igem.org/Part:BBa_M50060
https://parts.igem.org/Part:BBa_I746916
Tchounwou, Paul B et al. “Heavy Metals Toxicity and the Environment.” EXS 101 (2012): 133–164. PMC.
Mojtaba Taseidifar, Fatemeh Makavipour, Richard M. Pashley, A.F.M. Mokhlesur Rahman, Removal of heavy metal ions from water using ion flotation, Environmental Technology & Innovation, Volume 8, 2017, Pages 182-190, ISSN 2352-1864.
Manju Bhargavi Gumpu, Swaminathan Sethuraman, Uma Maheswari Krishnan, John Bosco Balaguru Rayappan, A review on detection of heavy metal ions in water – An electrochemical approach, Sensors and Actuators B: Chemical, Volume 213, 2015, Pages 515-533, ISSN 0925-4005.
Schwan, William R. “Regulation of fim Genes in Uropathogenic Escherichia Coli.” World journal of clinical infectious diseases 1.1 (2011): 17–25. PMC. Kline, Kimberly A.; Fälker, Stefan; Dahlberg, Sofia; Normark, Staffan; Henriques-Normark, Birgitta. "Bacterial Adhesins in Host-Microbe Interactions". Cell Host & Microbe. 5 (6): 580–592.
“Team:Harvard BioDesign.” Team:Freiburg/Project/Protein Purification, 2015.igem.org/Team:Harvard_BioDesign.
Kjærgaard, Kristian, Mark A. Schembri, and Per Klemm. “Novel Zn2+-Chelating Peptides Selected from a Fimbria-Displayed Random Peptide Library.” Applied and Environmental Microbiology 67.12 (2001): 5467–5473. PMC.
“Team:Exeter/Results.” Team:Waterloo/Parts, 2017.igem.org/Team:Exeter/Results.
Mustapha MU, Halimoon N (2015) Microorganisms and Biosorption of Heavy Metals in the Environment: A Review Paper. J Microb Biochem Technol 7:253-256.
Schembri, Mark A., and Per Klemm. “Heterobinary Adhesins Based on the Escherichia Coli FimH Fimbrial Protein.” Applied and Environmental Microbiology 64.5 (1998): 1628–1633.
Romantsov, T., Battle, AR., Hendel, JL., Martinac B., and JM Wood. “Protein localization in Escherichia coli cells: comparison of the cytoplasmic membrane proteins ProP, LacY, ProW, AqpZ, MscS, and MscL.” J Bacteriol. 2010 Feb;192(4):912-24.
Lindner, Ariel B., et al. “Asymmetric Segregation of Protein Aggregates Is Associated with Cellular Aging and Rejuvenation.” PNAS, National Academy of Sciences, 26 Feb. 2008, Rodríguez-Carmona, Escarlata, et al. “Isolation of Cell-Free Bacterial Inclusion Bodies.” Advances in Pediatrics., U.S. National Library of Medicine, 17 Sept. 2010;105 (8) 3076-3081