Computational protocol: The Core Components of Organelle Biogenesis and Membrane Transport in the Hydrogenosomes of Trichomonas vaginalis

Similar protocols

Protocol publication

[…] All of the identified protein sequences were manually annotated based on searches in the TrichDB (http://trichdb.org/trichdb/), Uniprot Protein knowledgebase (http://www.uniprot.org/), NCBI (http://blast.ncbi.nlm.nih.gov/Blast.cgi) and Pfam (http://pfam.sanger.ac.uk/search) databases. The protein sequences (<1000 residues) were submitted (i) against a 90% redundancy-reduced NCBI nr database by means of simple pair-wise alignment Psi-BLAST for 8 iterations at an e-value cutoff of 10−3 and (ii) against the Pfam 23.0 A+B database of families represented by multiple sequence alignments and hidden Markov models (HMMs) at an e-value of 0.044.TargetP and SignalP, based on the combination of artificial neural networks and hidden Markov models, respectively (http://www.cbs.dtu.dk/services/), together with PsortII (http://psort.ims.u-tokyo.ac.jp/), were used to predict the subcellular location.An application based on the NetBeans Platform (Hunter) was used to predict the subcellular localization of proteins according to their N-terminal amino acid sequence, as previously described . The following parameters were used: (i) the presequence start motif - ML[ACGQRSTV] or MS[ILV] or MIS or MTL; (ii) a cleavage site motif - R.F[TKILFSAGQ] or R[AFNESG][TYILFSAGQ] or K[AFNESG][TYILFSAGQ] or K.F[TKILFSAGQ]. The secondary structure and topology of alpha-helix integral membrane proteins was predicted using two bioinformatic tools: TMHMM (http://www.cbs.dtu.dk/services/) and Memsat3 (http://bioinf.cs.ucl.ac.uk/memsat/).To determine members of protein families, a hidden Markov model analysis was performed according to the method of Likic et al. . [...] The model of TvTim44 (residues 144–326) was built using the human Tim44 structure (PDB ID: 2cw9) as a template. The alignment was constructed by MUSCLE and manually edited. The 3D structure model of TvTim 44 was built using Modeller 9v7 . The quality of the final model was checked using What Check and ProSA-web services . The electrostatic potential on the solvent-accessible surface of TvTim44 was calculated using APBS 1.3 . […]

Pipeline specifications

Software tools MUSCLE, MODELLER, WHAT_CHECK, ProSA-web
Applications Protein structure analysis, Nucleotide sequence alignment
Organisms Trichomonas vaginalis, Saccharomyces cerevisiae
Diseases Trichomonas Infections
Chemicals Adenosine Diphosphate, Adenosine Triphosphate, Hydrogen