The FREAKS Session 3.1: Repeats Session 3.2: Biased regions Miguel Andrade Johannes-Gutenberg University of Mainz of PROTEIN SEQUENCE
Frequency 14% proteins contains repeats (Marcotte et al, 1999) 1: Single amino acid repeats. 2: Longer imperfect tandem repeats. Assemble in structure.
Definition repeats Sequence, long, imperfect, tandem MRAVVKSPIMCHEKSPSVCSPLNMTSSVCSPAGINSVSSTTASF GSFPVHSPITQGTPLTCSPNVENRGSRSHSPAHASNVGSPLSSP LSSMKSSISSPPSHCSVKSPVSSPNNVTLRSSVSSPANINN
Definition repeats Sequence, long, imperfect, tandem MRAVVKSPIMCHEKSPSVCSPLNMTSSVCSPAGINSVSSTTASF GSFPVHSPITQGTPLTCSPNVENRGSRSHSPAHASNVGSPLSSP LSSMKSSISSPPSHCSVKSPVSSPNNVTLRSSVSSPANINN
Definition repeats Sequence, long, imperfect, tandem MRAVVKSPIM CHE KSPSVCSPLN MTSSVCSPAG INSVSSTTASF GSFPVHSPIT Q GTPLTCSPNV EN RGSRSHSPAH ASN VGSPLSSPLS S MKSSISSPPS HCS VKSPVSSPNN VT LRSSVSSPAN INN
Definition repeats Sequence, long, imperfect, tandem MRAVVKSPIM CHE KSPSVCSPLN MTSSVCSPAG INSVSSTTASF GSFPVHSPIT Q GTPLTCSPNV EN RGSRSHSPAH ASN VGSPLSSPLS S MKSSISSPPS HCS VKSPVSSPNN VT LRSSVSSPAN INN
Tandem repeats fold together
Definition repeats Sequence, long, imperfect, tandem MRAVVKSPIM CHE KSPSVCSPLN MTSSVCSPAG INSVSSTTASF GSFPVHSPIT Q GTPLTCSPNV EN RGSRSHSPAH ASN VGSPLSSPLS S MKSSISSPPS HCS VKSPVSSPNN VT LRSSVSSPAN INN
(Vlassi et al, 2013)
Andrade et al. (2001) J Struct Biol
Definition CBRs Perfect repeat: QQQQQQQQQQQ Imperfect: QQQQPQQQQQQ Amino acid type: DDDDDEEEDEDEED Compositionally biased regions (CBRs) High frequency of one or two amino acids in a region. Particular case of low complexity region
Detection CBRs Sometimes straightforward. N-terminal human Huntingtin. How many CBRs can you find? >sp|P42858|HD_HUMAN Huntingtin OS=Homo sapiens MATLEKLMKAFESLKSFQQQQQQQQQQQQQQQQQQQQQPPPPPPPPPPPQLPQPPPQAQP LLPQPQPPPPPPPPPPGPAVAEEPLHRPKKELSATKKDRVNHCLTICENIVAQSVRNSPE FQKLLGIAMELFLLCSDDAESDVRMVADECLNKVIKALMDSNLPRLQLELYKEIKKNGAP RSLRAALWRFAELAHLVRPQKCRPYLVNLLPCLTRTSKRPEESVQETLAAAVPKIMASFG NFANDNEIKVLLKAFIANLKSSSPTIRRTAAGSAVSICQHSRRTQYFYSWLLNVLLGLLV PVEDEHSTLLILGVLLTLRYLVPLLQQQVKDTSLKGSFGVTRKEMEVSPSAEQLVQVYEL TLHHTQHQDHNVVTGALELLQQLFRTPPPELLQTLTAVGGIGQLTAAKEESGGRSRSGSI VELIAGGGSSCSPVLSRKQKGKVLLGEEEALEDDSESRSDVSSSALTASVKDEISGELAA SSGVSTPGSAGHDIITEQPRSQHTLQADSVDLASCDLTSSATDGDEEDILSHSSSQVSAV PSDPAMDLNDGTQASSPISDSSQTTTEGPDSAVTPSDSSEIVLDGTDNQYLGLQIGQPQD EDEEATGILPDEASEAFRNSSMALQQAHLLKNMSHCRQPSDSSVDKFVLRDEATEPGDQE NKPCRIKGDIGQSTDDDSAPLVHCVRLLSASFLLTGGKNVLVPDRDVRVSVKALALSCVG AAVALHPESFFSKLYKVPLDTTEYPEEQYVSDILNYIDHGDPQVRGATAILCGTLICSIL
Detection CBRs Sometimes straightforward. N-terminal human Huntingtin. How many CBRs can you find? >sp|P42858|HD_HUMAN Huntingtin OS=Homo sapiens MATLEKLMKAFESLKSFQQQQQQQQQQQQQQQQQQQQQPPPPPPPPPPPQLPQPPPQAQP LLPQPQPPPPPPPPPPGPAVAEEPLHRPKKELSATKKDRVNHCLTICENIVAQSVRNSPE FQKLLGIAMELFLLCSDDAESDVRMVADECLNKVIKALMDSNLPRLQLELYKEIKKNGAP RSLRAALWRFAELAHLVRPQKCRPYLVNLLPCLTRTSKRPEESVQETLAAAVPKIMASFG NFANDNEIKVLLKAFIANLKSSSPTIRRTAAGSAVSICQHSRRTQYFYSWLLNVLLGLLV PVEDEHSTLLILGVLLTLRYLVPLLQQQVKDTSLKGSFGVTRKEMEVSPSAEQLVQVYEL TLHHTQHQDHNVVTGALELLQQLFRTPPPELLQTLTAVGGIGQLTAAKEESGGRSRSGSI VELIAGGGSSCSPVLSRKQKGKVLLGEEEALEDDSESRSDVSSSALTASVKDEISGELAA SSGVSTPGSAGHDIITEQPRSQHTLQADSVDLASCDLTSSATDGDEEDILSHSSSQVSAV PSDPAMDLNDGTQASSPISDSSQTTTEGPDSAVTPSDSSEIVLDGTDNQYLGLQIGQPQD EDEEATGILPDEASEAFRNSSMALQQAHLLKNMSHCRQPSDSSVDKFVLRDEATEPGDQE NKPCRIKGDIGQSTDDDSAPLVHCVRLLSASFLLTGGKNVLVPDRDVRVSVKALALSCVG AAVALHPESFFSKLYKVPLDTTEYPEEQYVSDILNYIDHGDPQVRGATAILCGTLICSIL
Detection CBRs Sometimes straightforward. N-terminal human Huntingtin. How many CBRs can you find? >sp|P42858|HD_HUMAN Huntingtin OS=Homo sapiens MATLEKLMKAFESLKSFQQQQQQQQQQQQQQQQQQQQQPPPPPPPPPPPQLPQPPPQAQP LLPQPQPPPPPPPPPPGPAVAEEPLHRPKKELSATKKDRVNHCLTICENIVAQSVRNSPE FQKLLGIAMELFLLCSDDAESDVRMVADECLNKVIKALMDSNLPRLQLELYKEIKKNGAP RSLRAALWRFAELAHLVRPQKCRPYLVNLLPCLTRTSKRPEESVQETLAAAVPKIMASFG NFANDNEIKVLLKAFIANLKSSSPTIRRTAAGSAVSICQHSRRTQYFYSWLLNVLLGLLV PVEDEHSTLLILGVLLTLRYLVPLLQQQVKDTSLKGSFGVTRKEMEVSPSAEQLVQVYEL TLHHTQHQDHNVVTGALELLQQLFRTPPPELLQTLTAVGGIGQLTAAKEESGGRSRSGSI VELIAGGGSSCSPVLSRKQKGKVLLGEEEALEDDSESRSDVSSSALTASVKDEISGELAA SSGVSTPGSAGHDIITEQPRSQHTLQADSVDLASCDLTSSATDGDEEDILSHSSSQVSAV PSDPAMDLNDGTQASSPISDSSQTTTEGPDSAVTPSDSSEIVLDGTDNQYLGLQIGQPQD EDEEATGILPDEASEAFRNSSMALQQAHLLKNMSHCRQPSDSSVDKFVLRDEATEPGDQE NKPCRIKGDIGQSTDDDSAPLVHCVRLLSASFLLTGGKNVLVPDRDVRVSVKALALSCVG AAVALHPESFFSKLYKVPLDTTEYPEEQYVSDILNYIDHGDPQVRGATAILCGTLICSIL
Detection CBRs Sometimes straightforward. N-terminal human Huntingtin. How many CBRs can you find? >sp|P42858|HD_HUMAN Huntingtin OS=Homo sapiens MATLEKLMKAFESLKSFQQQQQQQQQQQQQQQQQQQQQPPPPPPPPPPPQLPQPPPQAQP LLPQPQPPPPPPPPPPGPAVAEEPLHRPKKELSATKKDRVNHCLTICENIVAQSVRNSPE FQKLLGIAMELFLLCSDDAESDVRMVADECLNKVIKALMDSNLPRLQLELYKEIKKNGAP RSLRAALWRFAELAHLVRPQKCRPYLVNLLPCLTRTSKRPEESVQETLAAAVPKIMASFG NFANDNEIKVLLKAFIANLKSSSPTIRRTAAGSAVSICQHSRRTQYFYSWLLNVLLGLLV PVEDEHSTLLILGVLLTLRYLVPLLQQQVKDTSLKGSFGVTRKEMEVSPSAEQLVQVYEL TLHHTQHQDHNVVTGALELLQQLFRTPPPELLQTLTAVGGIGQLTAAKEESGGRSRSGSI VELIAGGGSSCSPVLSRKQKGKVLLGEEEALEDDSESRSDVSSSALTASVKDEISGELAA SSGVSTPGSAGHDIITEQPRSQHTLQADSVDLASCDLTSSATDGDEEDILSHSSSQVSAV PSDPAMDLNDGTQASSPISDSSQTTTEGPDSAVTPSDSSEIVLDGTDNQYLGLQIGQPQD EDEEATGILPDEASEAFRNSSMALQQAHLLKNMSHCRQPSDSSVDKFVLRDEATEPGDQE NKPCRIKGDIGQSTDDDSAPLVHCVRLLSASFLLTGGKNVLVPDRDVRVSVKALALSCVG AAVALHPESFFSKLYKVPLDTTEYPEEQYVSDILNYIDHGDPQVRGATAILCGTLICSIL
Detection repeats Sometimes straightforward. N-terminal human Huntingtin. How many repeats can you find? >sp|P42858|HD_HUMAN Huntingtin OS=Homo sapiens MATLEKLMKAFESLKSFQQQQQQQQQQQQQQQQQQQQQPPPPPPPPPPPQLPQPPPQAQP LLPQPQPPPPPPPPPPGPAVAEEPLHRPKKELSATKKDRVNHCLTICENIVAQSVRNSPE FQKLLGIAMELFLLCSDDAESDVRMVADECLNKVIKALMDSNLPRLQLELYKEIKKNGAP RSLRAALWRFAELAHLVRPQKCRPYLVNLLPCLTRTSKRPEESVQETLAAAVPKIMASFG NFANDNEIKVLLKAFIANLKSSSPTIRRTAAGSAVSICQHSRRTQYFYSWLLNVLLGLLV PVEDEHSTLLILGVLLTLRYLVPLLQQQVKDTSLKGSFGVTRKEMEVSPSAEQLVQVYEL TLHHTQHQDHNVVTGALELLQQLFRTPPPELLQTLTAVGGIGQLTAAKEESGGRSRSGSI VELIAGGGSSCSPVLSRKQKGKVLLGEEEALEDDSESRSDVSSSALTASVKDEISGELAA SSGVSTPGSAGHDIITEQPRSQHTLQADSVDLASCDLTSSATDGDEEDILSHSSSQVSAV PSDPAMDLNDGTQASSPISDSSQTTTEGPDSAVTPSDSSEIVLDGTDNQYLGLQIGQPQD EDEEATGILPDEASEAFRNSSMALQQAHLLKNMSHCRQPSDSSVDKFVLRDEATEPGDQE NKPCRIKGDIGQSTDDDSAPLVHCVRLLSASFLLTGGKNVLVPDRDVRVSVKALALSCVG AAVALHPESFFSKLYKVPLDTTEYPEEQYVSDILNYIDHGDPQVRGATAILCGTLICSIL
Detection repeats Often NOT straightforward. N-terminal human Huntingtin. How many repeats can you find? >sp|P42858|HD_HUMAN Huntingtin OS=Homo sapiens MATLEKLMKAFESLKSFQQQQQQQQQQQQQQQQQQQQQPPPPPPPPPPPQLPQPPPQAQP LLPQPQPPPPPPPPPPGPAVAEEPLHRPKKELSATKKDRVNHCLTICENIVAQSVRNSPE FQKLLGIAMELFLLCSDDAESDVRMVADECLNKVIKALMDSNLPRLQLELYKEIKKNGAP RSLRAALWRFAELAHLVRPQKCRPYLVNLLPCLTRTSKRPEESVQETLAAAVPKIMASFG NFANDNEIKVLLKAFIANLKSSSPTIRRTAAGSAVSICQHSRRTQYFYSWLLNVLLGLLV PVEDEHSTLLILGVLLTLRYLVPLLQQQVKDTSLKGSFGVTRKEMEVSPSAEQLVQVYEL TLHHTQHQDHNVVTGALELLQQLFRTPPPELLQTLTAVGGIGQLTAAKEESGGRSRSGSI VELIAGGGSSCSPVLSRKQKGKVLLGEEEALEDDSESRSDVSSSALTASVKDEISGELAA SSGVSTPGSAGHDIITEQPRSQHTLQADSVDLASCDLTSSATDGDEEDILSHSSSQVSAV PSDPAMDLNDGTQASSPISDSSQTTTEGPDSAVTPSDSSEIVLDGTDNQYLGLQIGQPQD EDEEATGILPDEASEAFRNSSMALQQAHLLKNMSHCRQPSDSSVDKFVLRDEATEPGDQE NKPCRIKGDIGQSTDDDSAPLVHCVRLLSASFLLTGGKNVLVPDRDVRVSVKALALSCVG AAVALHPESFFSKLYKVPLDTTEYPEEQYVSDILNYIDHGDPQVRGATAILCGTLICSIL
Detection repeats Often NOT straightforward. N-terminal human Huntingtin. How many repeats can you find? EFQKLLGIAMELFLLCSDDAESDVRMVADECLNKVIKA CRPYLVNLLPCLTRTSKRP-EESVQETLAAAVPKIMAS NDNEIKVLLKAFIANLKSSSPTIRRTAAGSAVSICQHS TQYFYSWLLNVLLGLLVPVEDEHSTLLILGVLLTLRYL PSAEQLVQVYELTLHHTQHQDHNVVTGALELLQQLFRT
Detection repeats Often NOT straightforward. N-terminal human Huntingtin. How many repeats can you find? EFQKLLGIAMELFLLCSDDAESDVRMVADECLNKVIKA CRPYLVNLLPCLTRTSKRP-EESVQETLAAAVPKIMAS NDNEIKVLLKAFIANLKSSSPTIRRTAAGSAVSICQHS TQYFYSWLLNVLLGLLVPVEDEHSTLLILGVLLTLRYL PSAEQLVQVYELTLHHTQHQDHNVVTGALELLQQLFRT
The FREAKS Session 3.1: Repeats Session 3.2: Biased regions Miguel Andrade Johannes-Gutenberg University of Mainz of PROTEIN SEQUENCE
Frequency repeats Fraction of proteins annotated with the keyword REPEAT in SwissProt % Archaea 27/ Viruses 81/ Bacteria 299/ Fungi 232/ Viridiplantae 153/ Metazoa 1538/ Rest of Eukaryota 92/ (Andrade et al 2001)
Detection of repeats Dotplots Comparing a sequence against itself
Detection of repeats Dotplots TLRSSVSSPANINNS NMTSSVCSPANISV
Detection of repeats Dotplots TLRSSVSSPANINNS NMTSSVCSPANISV | 1 match
Detection of repeats Dotplots TLRSSVSSPANINNS NMTSSVCSPANISV ||| ||||| 8 matches
Detection of repeats Dotplots TLRSSVSSPANINNS NMTSSVCSPANISV | | 2 matches
Detection of repeats Dotplots TLRSSVSSPANINNS NMTSSVCSPANISV | 1 match
Detection of repeats Dotplots TLRSSVSSPANINNS NMTSSVCSPANISV 8
Detection of repeats Dotplots TLRSSVSSPANINNS NMTSSVCSPANISV 1821
Exercise 1
Obtain the sequence from UniProtsequence from UniProt Go to the Dotlet web pageDotlet Click on the input button and paste the sequence there Try to find combinations of parameters that show patterns in the dot plot Find repetitions clicking in the diagonal patterns Exercise 1. Using Dotlet with the human mineralocorticoid receptor (MR)
Detection of repeats Using a multiple sequence alignment helps Conserved repeated patterns JalView with Regular Expression searches
Detection of repeats Using a multiple sequence alignment helps Conserved repeated patterns JalView with Regular Expression searches
Detection of repeats Using a multiple sequence alignment helps Conserved repeated patterns JalView with Regular Expression searches
Detection of repeats Using a multiple sequence alignment helps Conserved repeated patterns JalView with Regular Expression searches Regular Expressions: [LS]P.A matches L or S, followed by P, followed by anything, followed by A
Detection of repeats Using a multiple sequence alignment helps Conserved repeated patterns JalView with Regular Expression searches Regular Expressions: [LS]P.A matches L or S, followed by P, followed by anything, followed by A Which one is not matched? LPTA, SPAA, LPPA, LPAP, SPLA
Detection of repeats Using a multiple sequence alignment helps Conserved repeated patterns JalView with Regular Expression searches Regular Expressions: [LS]P.A matches L or S, followed by P, followed by anything, followed by A Which one is not matched? LPTA, SPAA, LPPA, LPAP, SPLA
Run JalView using the JNLP file in desktop (from Load this MSA in JalViewthis MSA Use the "find" option with a regular expression and mark all matches Try to find the expression that matches more repeats. How many repeats do you see? How long are they? Would you correct the alignment based on these findings? Exercise 2. Using JalView with a MSA of the MR with orthologs
#T1 #T13#T12#T11#T10#T9#T8 #T7 #T2#T3#T4#T5#T6 #F1 #F2#F3 #F4 #F5#F10#F9#F8#F7#F6 #T14 #T15 #F11 *** **** (Vlassi et al, 2013)
Andrade and Bork (1995) Nature Genetics
A subunit PP2A structure PDB:1b3u Groves et al. (1999) Cell
Ap1 Clathrin Adaptor Core PDB:1w63 Heldwein et al. (2004) PNAS
Ap1 Clathrin Adaptor Core PDB:1w63 Heldwein et al. (2004) PNAS
Neural Network! Secondary structure Transmembrane helices Residue exposure Andrade, Petosa, O'Donoghue, Müller and Bork (2001) J Mol Biol
Neural Network Backpropagation neural network …GTAARTWCGASLFVPRLLAGHVDITSLVRALAKSGDLFVARSTKT… Central position Output neuron = Hidden layer n=3 Input layer n=39x20 Architecture Palidwor et al. (2009) PLoS Comp Biol
Neural Network Architecture H1H2 L H1H2L Fournier et al. (2013) PLoS One
Virus/phages E-05 Archaea E-04 Euryarchaeota E-03 Crenarchaeota E-04 Bacteria E-04 Acidobacteria E-04 Actinobacteria E-04 Bacteroidetes E-04 Chlamydiae E-03 Chloroflexi E-03 Cyanobacteria E-03 Firmicutes E-04 Planctomycetes E-03 Proteobacteria E-04 Spirochetes E-04 Eukaryota E-03 Apicomplexa E-03 Ciliophora E-03 Bacillariophyta E-03 Viridiplantae E-03 Chlorophyta E-03 Streptophyta E-03 Kinetoplastida E-03 Mycetozoa E-03 Phaeophyceae E-03 Fungi E-03 Metazoa E-03 Nematodes E-03 Trematodes E-03 Insecta E-03 Sarcopterygii E E-03 Fournier et al. (2013) PLoS One
Go to the ARD2 web pageARD2 Paste this sequence (1-780 fragment of human huntingtin) in the input windowthis sequence Run ARD2 and interpret the output Exercise 3. Detecting repeats in human huntingtin
Go to the PDBPaint web pagePDBPaint In the "Query PDB" window type "2IE3", in the "web service" menu choose the "ARD2" option and select a large window size (e.g. 800*800) Hit the "Go!" button Turn around the structure and examine the correspondence between the hits and the structure Exercise 4. Viewing detected repeats in a protein structure