Difference between revisions of "Part:BBa K808004"
 
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The large subunit A2 of the tripartite tricarboxylate transporter family (tctA_505, 52,99 kDa) was isolated from  ''Comamonas testosteroni KF-1.'' The tripartite tricarboxylate transporter system consists of three different proteins: a periplasmatic solute binding receptor, a membrane protein with 12 putative transmembrane alpha-helical spanners (in this case tctA_503), and a small poorly conserved membrane proteine with four putative transmembrane alpha-helical spanners<sup>[1]</sup>.The strain was purchased from Leibniz Institute DMSZ-German Collection of Microorganism and Cell Cultures (DMSZ no. 14576). The original sequence contains a Pst1 recognition site. To eliminate this recognition site a directed-site mutagenic PCR was performed. (For more datails: link zu dem PCR protokoll und dem Labjournal, wo isolations PCR beschrieben wird) To characterized the structure of the tctA_505 bioinformatic tools like Protein Homology/anologY Recognition Engine V 2.0 (PHYRE2), I-TASSER  servers, and TMHMM was used. The TMHMM predicted a  transmembrane protein with 11 alpha-helical spanners (Fig. 1).The NCBI Protein BLAST results shows that the tctA_505 subunit A2 belongs to the tctA superfamily.Phyre2 an I-Tasser server homology modelling did not give a significant result for the structure of the tctA_505 large subunit A2. ]]
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The large subunit A2 of the tripartite tricarboxylate transporter family (tctA_505, 52,99 kDa) was isolated from  ''Comamonas testosteroni KF-1.'' The tripartite tricarboxylate transporter system consists of three different proteins: a periplasmatic solute binding receptor, a membrane protein with 12 putative transmembrane alpha-helical spanners (in this case tctA_503), and a small poorly conserved membrane proteine with four putative transmembrane alpha-helical spanners<sup>[1]</sup>.The strain was purchased from Leibniz Institute DMSZ-German Collection of Microorganism and Cell Cultures (DMSZ no. 14576). The original sequence contains a Pst1 recognition site. To eliminate this recognition site a directed-site mutagenic PCR was performed. (For more details [http://2012.igem.org/Team:TU_Darmstadt/Protocols/mutagenic_PCR mutagenic PCR]). To characterized the structure of the tctA_505 bioinformatic tools like Protein Homology/anologY Recognition Engine V 2.0 (PHYRE2), I-TASSER  servers, and TMHMM was used. The TMHMM predicted a  transmembrane protein with 11 alpha-helical spanners (Fig. 1).The NCBI Protein BLAST results shows that the tctA_505 subunit A2 belongs to the tctA superfamily.Phyre2 an I-Tasser server homology modelling did not give a significant result for the structure of the tctA_505 large subunit A2. ]]
  
 
[[Image:File-tcta_tmhmm.png|900px|thumb|center|Figure 1. '''TMHMM prediction of the tctA_505 subunit A2.''' It shows 10 alpha-helical spanners.]]
 
[[Image:File-tcta_tmhmm.png|900px|thumb|center|Figure 1. '''TMHMM prediction of the tctA_505 subunit A2.''' It shows 10 alpha-helical spanners.]]

Latest revision as of 22:01, 26 September 2012

tctA_505: large subunit A2 of the tripartite tricarboxylate transporter family


The large subunit A2 of the tripartite tricarboxylate transporter family (tctA_505, 52,99 kDa) was isolated from Comamonas testosteroni KF-1. The tripartite tricarboxylate transporter system consists of three different proteins: a periplasmatic solute binding receptor, a membrane protein with 12 putative transmembrane alpha-helical spanners (in this case tctA_503), and a small poorly conserved membrane proteine with four putative transmembrane alpha-helical spanners[1].The strain was purchased from Leibniz Institute DMSZ-German Collection of Microorganism and Cell Cultures (DMSZ no. 14576). The original sequence contains a Pst1 recognition site. To eliminate this recognition site a directed-site mutagenic PCR was performed. (For more details [http://2012.igem.org/Team:TU_Darmstadt/Protocols/mutagenic_PCR mutagenic PCR]). To characterized the structure of the tctA_505 bioinformatic tools like Protein Homology/anologY Recognition Engine V 2.0 (PHYRE2), I-TASSER servers, and TMHMM was used. The TMHMM predicted a transmembrane protein with 11 alpha-helical spanners (Fig. 1).The NCBI Protein BLAST results shows that the tctA_505 subunit A2 belongs to the tctA superfamily.Phyre2 an I-Tasser server homology modelling did not give a significant result for the structure of the tctA_505 large subunit A2. ]]

Figure 1. TMHMM prediction of the tctA_505 subunit A2. It shows 10 alpha-helical spanners.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 557
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 193
    Illegal NgoMIV site found at 802
    Illegal NgoMIV site found at 1186
  • 1000
    COMPATIBLE WITH RFC[1000]


References

[1] Sasoh, M., E. Masai, et al. (2006). "Characterization of the terephthalate degradation genes of Comamonas sp. strain E6." Appl Environ Microbiol 72(3): 1825-1832.

  • Fukuhara, Y., K. Inakazu, et al. (2010). "Characterization of the isophthalate degradation genes of Comamonas sp. strain E6." Appl Environ Microbiol 76(2): 519-527.
  • Kamimura, N., T. Aoyama, et al. (2010). "Characterization of the protocatechuate 4,5-cleavage pathway operon in Comamonas sp. strain E6 and discovery of a novel pathway gene." Appl Environ Microbiol 76(24): 8093-8101.
  • Winnen, B., R. N. Hvorup, et al. (2003). "The tripartite tricarboxylate transporter (TTT) family." Res Microbiol 154(7): 457-465.
  • Protein structure prediction on the web: a case study using the PhyreKelley LA and Sternberg MJE.Nature Protocols 4, 363 - 371 (2009 server
  • Yang Zhang. I-TASSER server for protein 3D structure prediction. BMC Bioinformatics, vol 9, 40 (2008).
  • Ambrish Roy, Alper Kucukural, Yang Zhang. I-TASSER: a unified platform for automated protein structure and function prediction. Nature Protocols, vol 5, 725-738 (2010).
  • Ambrish Roy, Jianyi Yang, Yang Zhang. COFACTOR: An accurate comparative algorithm for structure-based protein function annotation. Nucleic Acids Research, doi:10.1093/nar/gks372 (2012)
  • Prediction of twin-arginine signal peptides. Jannick Dyrløv Bendtsen, Henrik Nielsen, David Widdick, Tracy Palmer and Søren Brunak. BMC bioinformatics 2005 6: 167.
  • SignalP 4.0: discriminating signal peptides from transmembrane regions Thomas Nordahl Petersen, Søren Brunak, Gunnar von Heijne & Henrik Nielsen Nature Methods, 8:785-786, 2011
  • Erik L.L. Sonnhammer, Gunnar von Heijne, and Anders Krogh:A hidden Markov model for predicting transmembrane helices in protein sequences. In Proc. of Sixth Int. Conf. on Intelligent Systems for Molecular Biology, p 175-182,Ed J. Glasgow, T. Littlejohn, F. Major, R. Lathrop, D. Sankoff, and C. Sensen Menlo Park, CA: AAAI Press, 1998