MOTIFS: USING DATABASES & CREATING YOUR OWN

SEARCHING MOTIF DATABASES

BACKGROUND INFORMATION:  Proteins having related functions may not show overall high homology yet may contain sequences of amino acid residues that are highly conserved. For background information on this see PROSITE at ExPASy. N.B. I recommend that you check your protein sequence with at least two different search engines.  Alternatively, use a meta site such as MOTIF (GenomeNet, Institute for Chemical Research, Kyoto University, Japan) to simultaneously carry out Prosite, Blocks, ProDom, Prints and Pfam search

Several great sites including the first four which are meta sites:

red_bullet.gif (914 bytes)  Motif Scan (MyHits, SIB, Switzerland) includes Prosite, Pfam and HAMAP profiles.
red_bullet.gif (914 bytes) InterPro 5 - includes PROSITE, HAMAP (High-quality Automated and Manual Annotation of Proteins), Pfam (protein Families), PRINTS, ProDom, SMART (a Simple Modular Architecture Research Tool), TIGRFAMs, PIRSF (Protein Information Resource), SUPERFAMILY, CATH-Gene3D (Class, Architecture, Topology, Homologous superfamily), and PANTHER  (Protein ANalysis THrough Evolutionary Relationships) classification systems. (Reference: Jones, P. et al. 2014, Bioinformatics 10: 1093)  . This service is also available here.

red_bullet.gif (914 bytes) MOTIF (GenomeNet, Japan) - I recommend this for the protein analysis, I have tried phage genomes against the DNA motif database without success. Offers 6 motif databases and the possibility of using your own.
red_bullet.gif (914 bytes) CDD or CD-Search (Conserved Domain Databases) - (NCBI) includes CDD, Smart,Pfam, PRK, TIGRFAM, COG and KOG  and is invoked when one uses BLASTP.  This tool can also be accessed here.

 Batch Web CD-Search Tool - The Batch CD-Search tool allows the computation and download of conserved domain annotation for large sets of protein queries. Input up to 100,000 protein query sequences as a list of sequence identifiers and/or raw sequence data, then download output in a variety of formats (including tab-delimited text files) or view the search results graphicallyOn the Batch CD-Search job summary page, a "Browse Results" button above the sample data table allows you to view the results graphically. The button opens a separate browser window that shows the domain footprints, alignment details, and conserved features on any individual query sequence..(Reference: Marchler-Bauer A et al. 2011. Nucleic Acids Res.39: (D)225-229.)

red_bullet.gif (914 bytes) Pfam  - (Sanger Institute) while for Batch Pfam searc hes go here.

red_bullet.gif (914 bytes) ScanProsite (ExPASy)
red_bullet.gif (914 bytes) Block Searcher (Fred Hutchinson Cancer Research Center, U.S.A.)
red_bullet.gif (914 bytes) PRINTS (University of Manchester, UK) - is a compendium of protein fingerprints. A fingerprint is a group of conserved motifs used to characterise a protein family; its diagnostic power is refined by iterative scanning of a SWISS-PROT/TrEMBL composite. Usually the motifs do not overlap, but are separated along a sequence, though they may be contiguous in 3D-space. Fingerprints can encode protein folds and functionalities more flexibly and powerfully than can single motifs, full diagnostic potency deriving from the mutual context provided by motif neighbours. For Scan use against PRINTS use  P-val FingerPRINTScan or PRINTS Blast .


ProDom (Pôle Rhone-Alpin de BioInformatique, France ) - is a comprehensive set of protein domain families automatically generated from the UniProt Knowledge Database
SMART Simple Modular Architecture Research Tool (EMBL, Universitat Heidelberg) - searches sequence for the domains/ sequences listed in the homepage.  Try selecting/deselecting the default settings.

 Batch SMART scan - can be found  herePlease note that the software produces a polyprotein which it analyzes.  This can result in some difficulty in correlating the motifs which the individual proteins.  The same proviso applies to the Batch CD search.

iProClass (Protein Information Resource, Georgetown University Medical Centre, U.S.A.) - is an integrated resource that provides comprehensive family relationships and structural/functional features of proteins.

red_bullet.gif (914 bytes) MEROPS BLAST - permits one to screen protein sequences against an extensive database of characterized peptidases (Rawlings, N.D., O'Brien, E. A. & Barrett, A.J. (2002) MEROPS: the protease database. Nucleic Acids Res. 30, 343-346).

 MEROPS batch BLAST - can be accessed here.

For specific protein modifications or site detection consult the following sites:

DNA binding - motifs: (A good tutorial resource can be found here )

 GYM - the most recent program for analysis of helix-turn-helix motifs in proteins.  N.B. the next site dates from 1990. (Reference: Narasimhan, G. et al. 2002. J. Computational Biol. 9:707-720)
 Helix-turn-Helix Motif Prediction - (Institut de Biologie et Chemie des Proteines, Lyon, France)

 iDNA-Prot - identifies DNA-binding proteins via the “grey model” and by adopting the random forest operation engine.The overall success rate by iDNA-Prot was 83.96%. One can submit up to 50 proteins. (Reference: Lin W-Z et al. 2011. PLoS One 6: e24756). Also available here.

  HTHQuery - is another site for detection of the DNA-binding helix-turn-helix motif. This program has a true positive rate of 83.5% and a false positive rate of 0.8%. Unfortunately it only takes a pdb file. (Reference: C. Ferrer-Costa et al. 2005. Bioinformatics 2005 21: 3679-3680).
  DNAbinder - employs two approaches to predict DNA-binding proteins (a) amino acid composition which allows for multiple sequences in fasta format, and (b) PSSM (Position-specific scoring matrix) which can only screen a single protein at a time.  Choose the "Alternate dataset" if input sequence is full length protein, since the prediction will be done using SVM modules developed using full length protein sequences (Reference: M. Kumar et al. 2007. BMC Bioinformatics 8: 463).
  DNABindR - uses a "Naive Bayes" prediction model to predict protein-DNA interaction sites (Reference: C. Yan  et al. 2006. BMC Bioinformatics 7:262)
  BindN - applies support vector machines (SVMs) to prediction of DNA and RNA-binding residues from sequence features, including the side chain pKa value, hydrophobicity index and molecular mass of an amino acid (Reference: Wang, L. and Brown, S.J.. 2006. Nucl. Acids Res. 34: W243-248).
  DP-Bind - prediction can be performed using a profile of evolutionary conservation of the input sequence automatically generated by the web-server or the input sequence alone. Three prediction methods are run for each input sequence and consensus prediction is generated (Reference: S. Hwang et al. 2007. Bioinformatics 25: 634-636).

If you know the three-dimensional structure of your protein then 3D-footprint, DISPLAR (Reference: Tjong G & Zhou H-X. 2007. Nucl. Acid Res.35: 1465-1477), iDBPs (Reference: Nimrod G. et al. 2009. J. Mol. Biol. 387: 1040-1053), DNABIND (Reference: Szlagyi A & Skolnick J. 2006. J. Mol. Biol. 358: 922-933); and, DNABINDPROT (Reference: Ozbek P et al. 2010. Nucl. Acids Res. 38: W417-423) could be useful to you..

  2ZIP - is used to find leucine zipper motifs (Reference:   Bornberg-Bauer,E. et al. (1998) Nucleic Acids Res. 26:2740-2746).

 BCPREDS server allows users to choose the method for predicting B-cell epitopes among several developed prediction methods: AAP method, BCPred and FBCPred. Users provide an antigen sequence and optionally can specify desired epitope length and specificity threshold. Results are returned in several user-friendly formats. (Reference: EL-Manzalawy, Y. et al. 2008. J Mol Recognit 21: 243-255)

 EpiSearch: Mapping of Conformational Epitopes (Reference: Negi, S.S. & Braun, W. 2009. Bioinform. Biol. Insights 3: 71-81).

 CEP - Conformational Epitope Prediction Server - The algorithm, apart from predicting conformational epitopes, also predicts antigenic determinants and sequential epi-topes. The epitopes are predicted using 3D structure data of protein antigens, which can be visualized graphically. The algorithm employs structure-based Bioinformatics approach and solvent accessibility of amino acids in an explicit manner. Accuracy of the algorithm was found to be 75% when evaluated using X-ray crystal structures of Ag–Ab complexes available in the PDB.(Reference: Kulkarni-Kale, U. et al. 2005. Nucl. Acids Res. 33: W168–W171)

 SEEPA (Reference: Sun, J. 2009. Nucleic Acids Res. 37(Web Server issue):W612-6).

Glycosylation:

NetOGlyc (Center for Biological Sequence Analysis, Technical University of Denmark) - produces neural network predictions of mucin type GalNAc O-glycosylation sites in mammalian proteins. SignalP is automatically run on all sequences. A warning is displayed if a signal peptide is not detected. In transmembrane proteins, only extracellular domains may be O-glycosylated with mucin-type GalNAc.
NetNGlyc (Center for Biological Sequence Analysis, Technical University of Denmark) - predicts N-Glycosylation sites in human proteins using artificial neural networks that examine the sequence context of Asn-Xaa-Ser /Thr sequons.
YinOYang (Center for Biological Sequence Analysis, Technical University of Denmark) - produces neural network predictions for O-ß-GlcNAc attachment sites in eukaryotic protein sequences. This server can also use NetPhos, to mark possible phosphorylated sites and hence identify "Yin-Yang" sites.

Fatty acylation:

LipoP 1.0 (Center for Biological Sequence Analysis Technical University of Denmark) - allows prediction of where signal peptidases I & II cleavage sites from Gram negative bacteria will cleave a protein. 

NMT - The MYR Predictor (IMP [Research Institute of Molecular Pathology] Bioinformatics Group, Austria) - predicts N-terminal N-myristoylation. Generally, the enzyme NMT requires an N-terminal glycine (leading methionines are cleaved prior to myristoylation). However, also internal glycines may become N-terminal as a result of proteolytic processing of proproteins.
Myristoylator (ExPASy, Switzerland) - predicts N-terminal myristoylation of proteins by neural networks. Only N-terminal glycines are myristoylated (leading methionines are cleaved prior to myristoylation).

Nucleotide binding sites:

 nSITEpred - is designed for sequence-based prediction of binding residues for ATP, ADP, AMP, GDP, and GTP (Reference: K. Chen 2012. Bioinformatics 28: 331-341)

Phosphorylation:

GPS (Group-based Phosphorylation Scoring method) - prediction encompases 71 Protein Kinase (PK) families/PK groups (Reference: Y. Xue et al. 2005. Nucl. Acids Res. 33: W184-W187).

KinasePhos - this method is purported to have higher accuracy and provides not only the location of the phosphorylation sites, but also the corresponding catalytic protein kinases. (Reference: H.-D. Huang et al. 2005. Nucl. Acids Res. 33: W226-W229).

NetPhos (Center for Biological Sequence Analysis, Technical University of Denmark) - predicts Ser, Thr and Tyr phosphorylation sites in eukaryotic proteins.

Sulfation:

The Sulfinator  (ExPASy, Switzerland) predicts tyrosine sulfation sites in protein sequences.  

DISCOVER YOUR OWN MOTIFS: 

After you have discovered similar sequences but the motif searching tools have failed to recognize your group of proteins you can use the following tools to create a list of potential motifs.

red_bullet.gif (914 bytes) The MEME Suite- Motif-based sequence analysis tools    (National Biomedical Computation Resource, U.S.A.). N.B. After doing a BLASTP search create a FASTA-formated document containing three or four of the most homologous proteins (training set) and submit to MEME ( Multiple EM for Motif Elicitation) or GLAM2 (Gapped Local Alignments of Motifs).  In the case of MEME I usually specify 5 as the "Maximum number of motifs" to find. You will receive a message by E-mail entitled "MEME Submission Information (job app......)," verifies that the NBCR received and is processing your request.  If you click on the hyperlink "You can view your job results at: http://meme..." you will see:

The "MAST output as HTML" provides the motifs, a motif alignment graphic and the alignment of the motifs with the individual sequences in the training set. The "MEME output as HTML" file contains a detailed analysis of each of the motifs plus their Sequence Logos. 

At the top of the life is a buttom labelled "Search sequence databases for the best combined matches with these motifs using MAST."  This will take you to theMAST (Motif Alignment and Search Tool) submission form. Click on the NCBI nonredundant protein database.  You will receive an E-mail entitled "MAST Submission Information (job app ...)."

Use great caution before printing the second set of data can be >20 pages (Reference: Bailey, T.L. et al. 2009. Nucl. Acids Res. 37(Web Server issue): W202-W208).  There are two mirror sites for the Meme Suite - here  and here.

red_bullet.gif (914 bytes) Block Maker - (Fred Hutchinson Cancer Research Center, U.S.A.) Block Maker finds conserved blocks in a group of two or more unaligned related protein sequences. One can use the motifs to in data base searches, to construct trees and web logos.

red_bullet.gif (914 bytes) WebLogoa great graphical way of representing and visualizing consensus sequence data developed by Tom Schneider and Mike Stephens.  For nucleotide logos see RNA Structure Logo (The Technical University of Denmark)

red_bullet.gif (914 bytes) Seq2Logo is a sequence logo generator. Sequence logos are a graphical representation of the information content stored in a multiple sequence alignment (MSA) and provide a compact and highly intuitive representation of the position-specific amino acid composition of binding motifs, active sites, etc. in biological sequences.(Reference: Thomsen, M.C., & Nielsen, M. 2012. Nucleic Acids Res. 40(Web Server issue):W281-287).

red_bullet.gif (914 bytes) Skylign is a tool for creating logos representing both sequence alignments and profile hidden Markov models. Submit to the form in order to produce (i) interactive logos for inclusion in webpages, or (ii) static logos for use in documents.  Skylign accepts sequence alignments in any format accepted by HMMER (this includes Stockholm and aligned fasta format). (Reference: Wheeler TJ, et al. 2014. BMC Bioinformatics. 15: 7.).  The HMMER-formatted profile HMM files can be generated from an *.aln ClustalW file by pasting your ClustalW alignment (& title) into HMMBUILD (Pôle Bioinformatique Lyonnais, France) and use the output (saved as a *.hmm file) at Skylign.

red_bullet.gif (914 bytes) Two Sample Logo - detects and displays statistically significant differences in position-specific symbol compositions between two sets of multiple sequence alignments. In a typical scenario, two groups of aligned sequences will share a common motif but will differ in their functional annotation. Also available as a Java tool. (Reference: Vacic, V. et al. 2006. Bioinformatics 22: 1536-1537)

NUCLEIC ACID MOTIFS: (See also here)

red_bullet.gif (914 bytes) Rfam (Welcome Trust Sanger Institute, England) - permits one to analyze 2 kb of DNA for 36 structural or functional RNAs such as 5S rRNA, tRNA, tmRNA, group I & II catalytic introns, hammerhead ribozymes, signal recognition particles.