Difference between revisions of "Part:BBa K3843000:Design"

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===Design Notes===
 
===Design Notes===
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The GenBank sequence coded for a propeptide that also contained a signal sequence. We first codon optimized the sequence for Escherichia coli K12 and simultaneously removed illegal restriction sites for RFC[25]. Next, we identified the signal peptide using the bioinformatics server SignalP 5.0, then we removed the signal peptide. We also removed the last 142 bp from the sequence, as these were annotated to not be present in the mature peptide by the authors of the GenBank submission (Male et al., 2005). The resulting sequence gives rise to functional salmon trypsin, appropriate for use in a fusion protein.
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The GenBank sequence coded for a propeptide that also contained a signal sequence. We first codon optimized the sequence for <i>Escherichia coli K12</i> and simultaneously removed illegal restriction sites for RFC[25]. Next, we identified the signal peptide using the bioinformatics server SignalP 5.0, then we removed the signal peptide. We also removed the last 142 bp from the sequence, as these were annotated to not be present in the mature peptide by the authors of the GenBank submission (Male et al., 2005). The resulting sequence gives rise to functional salmon trypsin, appropriate for use in a fusion protein.
  
 
===Source===
 
===Source===
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Male, R., Lorens, J. B., Smalås, A. O., & Torrissen, K. R. (2005, March 3). Molecular cloning and characterization of anionic and cationic variants of trypsin from Atlantic Salmon. <i>European Journal of Biochemistry, 232</i>(2), 677-685. https://febs.onlinelibrary.wiley.com/doi/full/10.1111/j.1432-1033.1995.677zz.x?sid=nlm%3Apubmed.
 
Male, R., Lorens, J. B., Smalås, A. O., & Torrissen, K. R. (2005, March 3). Molecular cloning and characterization of anionic and cationic variants of trypsin from Atlantic Salmon. <i>European Journal of Biochemistry, 232</i>(2), 677-685. https://febs.onlinelibrary.wiley.com/doi/full/10.1111/j.1432-1033.1995.677zz.x?sid=nlm%3Apubmed.
 +
 +
Pettersen, E. F.; Goddard, T. D.; Huang, C. C.; Couch, G. S.; Greenblatt, D. M.; Meng, E. C. & Ferrin, T. E. UCSF Chimera--a visualization system for exploratory research and analysis (Version 1.15). <i>J Comput Chem</i>. 2004; <i>25</i>(13): 1605-1612. https://www.ncbi.nlm.nih.gov/pubmed/15264254
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RCSB PDB. (n.d.). <i>1UTM</i>. Retrieved October 17, 2021, from https://www.rcsb.org/structure/1UTM

Latest revision as of 18:01, 19 October 2021


PEA-binding salmon trypsin (1UTM)


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 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]


Design Notes

The GenBank sequence coded for a propeptide that also contained a signal sequence. We first codon optimized the sequence for Escherichia coli K12 and simultaneously removed illegal restriction sites for RFC[25]. Next, we identified the signal peptide using the bioinformatics server SignalP 5.0, then we removed the signal peptide. We also removed the last 142 bp from the sequence, as these were annotated to not be present in the mature peptide by the authors of the GenBank submission (Male et al., 2005). The resulting sequence gives rise to functional salmon trypsin, appropriate for use in a fusion protein.

Source

The sequence of this part was obtained from GenBank: https://www.ncbi.nlm.nih.gov/nuccore/X70071.

References

Leiros, H. K. S., Brandsdal, B. O., Andersen, O. A., Os, V., Leiros, I., Helland, R., Otlewski, J., Willassen, N. P., & Smalås, A. O. (2009, January 1). Trypsin specificity as elucidated by lie calculations, x‐ray structures, and association constant measurements. Protein Science, 13(4), 1056-1070. https://onlinelibrary.wiley.com/doi/full/10.1110/ps.03498604.

Male, R., Lorens, J. B., Smalås, A. O., & Torrissen, K. R. (2005, March 3). Molecular cloning and characterization of anionic and cationic variants of trypsin from Atlantic Salmon. European Journal of Biochemistry, 232(2), 677-685. https://febs.onlinelibrary.wiley.com/doi/full/10.1111/j.1432-1033.1995.677zz.x?sid=nlm%3Apubmed.

Pettersen, E. F.; Goddard, T. D.; Huang, C. C.; Couch, G. S.; Greenblatt, D. M.; Meng, E. C. & Ferrin, T. E. UCSF Chimera--a visualization system for exploratory research and analysis (Version 1.15). J Comput Chem. 2004; 25(13): 1605-1612. https://www.ncbi.nlm.nih.gov/pubmed/15264254

RCSB PDB. (n.d.). 1UTM. Retrieved October 17, 2021, from https://www.rcsb.org/structure/1UTM