Developing and Using Special Developing and Using Special Developing and Using Special Purpose Hidden Markov Model Purpose Hidden Markov Model Purpose Hidden Markov Model Databases Databases Databases Martin Gollery Associate Director of Bioinformatics University of Nevada, Reno Mgollery@unr.edu
Today’ ’s Tutorial s Tutorial Today • Instructor: Martin Gollery • Associate Director of Bioinformatics, University of Nevada, Reno • Consultant to several organizations • Formerly with TimeLogic • Developed several HMM databases
Hidden Markov Models Hidden Markov Models • What HMM’s are • Which HMM programs are commonly used • What HMM databases are available • Why you would use one DB over another • Integrated Resources- InterPro and more • How you can build your own HMM DB • Problems with building your own • Live demonstration
Hidden Markov Models- - Hidden Markov Models What are they, anyway? What are they, anyway? • Statistical description of a protein family's consensus sequence • Conserved regions receive highest scores • Can be seen as a Finite State Machine
Representation of Family Representation of Family Members Members yciH KDGII • ZyciH KDGVI • VCA0570 KDGDI • HI1225 KNGII • sll0546 KEDCV • C D E G I K N V 1 1.0 2 0.6 0.2 0.2 3 0.2 0.8 4 0.2 0.2 0.4 0.2 5 0.8 0.2
Representation of gaps in Family Representation of gaps in Family Members Members yciH KDGII • ZyciH KDGVI • VCA0570 KDGDI • HI1225 KNGII • sll0546 KED-V • C D E G I K N V - 1 1.0 2 0.6 0.2 0.2 3 0.2 0.8 4 0.2 0.4 0.2 0.2 5 0.8 0.2
For Maximum sensitivity- - For Maximum sensitivity C D E G I K N V - 1 1.0 2 0.6 0.2 0.2 3 0.2 0.8 4 0.2 0.4 0.2 0.2 5 0.8 0.2 No residue at any position should have a zero probability, even if it was not seen in the training data.
Start with an MSA… … Start with an MSA CLUSTAL W (1.7) multiple sequence alignment • yciH KDGVIEIQGDKRDLLKSLLEAKGMKVKLAGG • ZyciH KDGVIEIQGDKRDLLKSLLEAKGMKVKLAGG • VCA0570 KDGDIEIQGDVRDQLKTLLESKGHKVKLAGG • HI1225 KNGIIEIQGEKRDLLKQLLEQKGFKVKLSGG • sll0546 KEDCVEIQGDQREKILAYLLKQGYKAKISGG • PA4840 KDGVVEIQGEHVELLIDELLKRGFKAKKSGG • AF0914 KNGVIELQGNHVNRVKELLIKKGFNPERIKT • *:. :*:**: : : * :* : : •
Hidden Markov Models Hidden Markov Models HMMER2.0 • NAME example2 • DESC Small example for demonstration purposes • LENG 31 • ALPH Amino • COM hmmbuild example2 example2.aln • NSEQ 7 • DATE Wed Jan 08 13:33:06 2003 • HMM A C D E F G H I K … • 1 -3217 -3413 -3082 -2664 -4291 -3257 -2104 -4231 3883 … • 2 -1938 -3859 2747 1592 -4024 -1857 -1206 -3953 -1455 … • 3 -2160 -3144 1834 -953 -4284 3247 -2013 -4362 -2365 … • 4 -1255 2750 436 -2789 -1273 -2972 -2049 1510 -2543 … • 5 -2035 -1558 -4660 -4320 -2085 -4409 -4229 3081 -4224 … • 6 -3264 -3765 -1447 3822 -4535 -2948 -2636 -4814 -2810 … • 7 -2423 -1951 -4843 -4395 -1156 -4544 -3680 3291 -4151 … • 8 -3220 -3396 -2530 -2667 -3851 -3171 -2735 -4442 -2277 … • 9 -3196 -3194 -3915 -4259 -4867 3789 -4005 -5414 -4591 … • 10 -1923 -3837 2743 2134 -4005 -1854 -1196 -3929 -1434 … • 11 -999 -2164 -952 -353 -2483 -1909 3321 -2139 1730 … • 12 -1629 -1909 -2827 -2102 -2279 -2588 -1442 -1012 -488 … •
Emission Probabilities Emission Probabilities • What is the likelihood that sequence X was emitted by HMM Y? • Likelihood is calculated by adding the probability of each residue at each position, and each of the transition probabilities
Plan7 from Outer Space Plan7 from Outer Space (Well, from St. Louis, anyway!) (Well, from St. Louis, anyway!)
HMM’ ’s s vs BLAST vs BLAST HMM • Position specific scoring vs. general matrix • Example: dDGVIvIddDKRDLLKSLiEAKkMKVKLAGG – KDGVIEIQGDKRDLLKSLLEAKGMKVKLAGG has 80% BLAST – similarity, but misses highly conserved regions • Scoring emphasizes important locations • Clearer score cutoffs • However, it is MUCH slower!
HMM programs HMM programs • HMMer -Sean Eddy, Wash U • SAM - Haussler, UCSC • Wise tools - Birney, EBI • SledgeHMMer - Subramaniam, SDSC • Meta-MEME - Noble & Bailey • PSI-BLAST - NCBI • SPSpfam - Southwest Parallel Software • Ldhmmer - Logical Depth • DeCypherHMM - TimeLogic
What exactly do you want? What exactly do you want? • Are you searching thousands of sequences with one or a few models? • Use hmmsearch • Searching a few sequences with thousands of models? • Use hmmpfam • Thousands of sequences vs. Thousands of models? • Use an accelerator, if you do it very often
HMM databases HMM databases • PFAM • TIGRFAM • Superfamily • SMART • Panther • PRED-GPCR
HMM databases at the CFB HMM databases at the CFB • COGfam • KinFam • HydroHMMer • NVfam-pro • NVfam-arc • NVfam-fun • NVfam-pln
PFAM PFAM • From Sanger, WashU, KI, INRA • Version 17 has 7868 families • Most widely used HMM database • Good annotation team
PFAM PFAM • PFAM-A is hand curated • From high quality multiple Alignments • PFAM-B is built automatically from ProDom • Generated using the Domainer algorithm • ProDom is built from SP/TREMBL
PFAM PFAM • Pfam-ls = global alignments • Pfam-fs = local alignments, so that matches may include only part of the model • Both the –ls and –fs versions are local W.R.T. the sequence
PFAM PFAM • Note ‘type’ annotation • Labeled TP • Family • Domain • Repeat • Motif
TIGRFAMs TIGRFAMs • Available at (www.tigr.org/TIGRFAMs/) • Organized by functional role • Equivalogs: a set of homologous proteins that are conserved with respect to function since their last common ancestor • Equivalog domains: domains of conserved function
TIGRFAMs TIGRFAMs • 2453 models in release 4.1 • Complementary to PFAM, so run both • Part of the Comprehensive Microbial Resource (CMR)
TIGRFAMs TIGRFAMs TIGRfam and PFAM alignments for Pyruvate carboxylase. The thin line represents the sequence. The bars represent hit regions.
SuperFamily SuperFamily • By Julian Gough, formerly MRC, now Riken GSC • www.supfam.org • Provides structural (and hence implied functional) assignments to protein sequences at the superfamily level • Built from SCOP (Structural Classification of Proteins) database, which is built from PDB • Available in HMMer, SAM, and PSI-BLAST formats
SuperFamily SuperFamily • 1447 SCOP Superfamilies • Each represented by a group of HMMs • Over 8500 models total • Table provides comparison to GO, Interpro, PFAM
SMART SMART • Simple Modular Architecture Research Tool • Version 3.4 contains 654 HMMs • Emphasis on mobile eukaryotic domains • smart.embl-heidelberg.de • Annotated with respect to phyletic distributions, functional class, tertiary structures and functionally important residues
SMART SMART • Use for signaling domains or extracellular domains • Normal and Genomic mode
PRED- -GPCR GPCR PRED • Papasaikas et al, U of Athens • 265 HMMs in 67 GPCR families • Based on TiPs Pharmacological classification. • Filters with CAST • signatures regularly updated • Entire system redone each year
-GPCR webserver GPCR webserver PRED- PRED
Panther Panther • Protein ANalysis THrough Evolutionary Relationships • Family and subfamily: families are evolutionarily related proteins; subfamilies are related proteins with the same function • Molecular function: the function of the protein by itself or with directly interacting proteins at a biochemical level, e.g. a protein kinase • Biological process: the function of the protein in the context of a larger network of proteins that interact to accomplish a process at the level of the cell or organism, e.g. mitosis. • Pathway: similar to biological process, but a pathway also explicitly specifies the relationships between the interacting molecules.
Panther Panther • (Thomas et al., Genome Research 2003; Mi et al. NAR 2005) • 6683 protein families • 31,705 functionally distinct protein subfamilies.
Panther Panther • Due to the size, searches could be slow • First, BLAST against consensus seqs • Then, search against models represented by those hits • With an accelerator, you don’t have to do that…
Panther Panther • So- how does it perform? • I took 3451 Arabidopsis proteins with no hit to PFAM, Superfamily, SMART or TIGRfam • Ran it against Panther • Found 160 significant hits!
COG- -HMMs HMMs COG • Clusters of Orthologous Groups of proteins • www.ncbi.nlm.nih.gov/cog/ • Each COG is from at least 3 lineages • Ancient conserved domain • 4873 alignments available • Alignments from NCBI, HMMs from me at mgollery@unr.edu
CDD CDD • Conserved Domain Database (NCBI) • Psi-BLAST profiles are similar to HMMs • 10991 PSSMs - SMART + COG +KOG+ Pfam+CD • Runs with RPS-BLAST • Much faster searches
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