Download Deduction and Visualization of Protein Structures

Deduction and
Visualization of Protein
Structures
Presented by Soodtida
Tangpraphaphorn
Protein Basics
 Amino acids have different charges
and polarities
 Primary structure is the sequence of
amino acids
 Secondary structure is the
interactions between a.a. side
chains
 Tertiary structure is the folded
conformation of a polypeptide
 Quaternary structure is the
interaction of polypeptide subunits
Applying Methods
1. Sequence the gene
2. Determine primary structure
(a.a. sequence)
3. Search for motifs, regions of
hydrophobicity/hydrophilicity,
salt bridges, sulfur bridges
4. Find similar polypeptides
5. Attempt to visualize structure
6. Speculate function if possible
Our Gene of Interest
1 atgcagttcg tgaacaagca gttcaactat aaggaccctg taaacggtgt tgacattgcc 61 tacatcaaaa ttccaaacgc
cggccagatg cagccggtga aggctttcaa gattcataac 121 aaaatctggg ttattccgga acgcgataca tttacgaacc
cggaagaagg agacttgaac 181 ccgccgccgg aagcaaagca ggtgccagtt tcatactacg attcaaccta tctgagcaca 241
gacaacgaga aggataacta cctgaaggga gtgaccaaat tattcgagcg tatttattcc 301 actgacctgg gccgtatgct
gctgacctca atcgtccgcg gaatcccatt ttggggtggc 361 agtaccattg acacggagtt gaaggttatt gacactaact
gcattaacgt gatccaacca 421 gacggtagct acagatctga agaacttaac ctcgtaatca tcgggccctc cgcggacatt 481
atccagtttg agtgcaagag ctttggccac gaagtgttga acctgacgcg taacggttac 541 ggctctactc agtacattcg
tttcagccca gacttcacgt tcggtttcga ggagagcctg 601 gaggttgata ccaacccgct gttgggtgca ggcaagttcg
caactgatcc agcggtgacc 661 ctggcacacg agctgatcca cgccggtcat cgtctgtatg gcattgcgat taacccgaac 721
cgcgtgttca aggttaacac caacgcctac tacgagatga gtggtttaga agtaagcttc 781 gaggaactgc gcacgttcgg
tggccatgat gcgaagttta tcgacagctt gcaggagaac 841 gagttccgtc tgtactacta caacaagttt aaagatattg
caagtacact gaacaaggct 901 aagtccattg tgggtaccac tgcttcatta cagtatatga aaaatgtttt taaagagaaa 961
tatctcctat ctgaagatac atctggaaaa ttttcggtag ataaattaaa atttgataag 1021 ttatacaaaa tgttaacaga gatttacaca
gaggataatt ttgttaagtt ttttaaagta 1081 cttaacagaa aaacatattt gaattttgat aaagccgtat ttaagataaa tatagtacct
1141 aaggtaaatt acacaatata tgatggattt aatttaagaa atacaaattt agcagcaaac 1201 tttaatggtc aaaatacaga
aattaataat atgaatttta ctaaactaaa aaattttact 1261 ggattgtttg aattttataa gttgctatgt gtaagaggga taataacttc
taaaactaaa 1321 tcattagata aaggatacaa taaggcatta aatgatttat gtatcaaagt taataattgg 1381 gacttgtttt
ttagtccttc agaagataat tttactaatg atctaaataa aggagaagaa 1441 attacatctg atactaatat agaagcagca
gaagaaaata ttagtttaga tttaatacaa 1501 caatattatt taacctttaa ttttgataat gaacctgaaa atatttcaat agaaaatctt
1561 tcaagtgaca ttataggcca attagaactt atgcctaata tagaaagatt tcctaatgga 1621 aaaaagtatg agttagataa
atatactatg ttccattatc ttcgtgctca agaatttgaa 1681 catggtaaat ctaggattgc tttaacaaat tctgttaacg aagcattatt
aaatcctagt 1741 cgtgtttata catttttttc ttcagactat gtaaagaaag ttaataaagc tacggaggca 1801 gctatgtttt
taggctgggt agaacaatta gtatatgatt ttaccgatga aactagcgaa 1861 gtaagtacta cggataaaat tgcggatata
actataatta ttccatatat aggacctgct 1921 ttaaatatag gtaatatgtt atataaagat gattttgtag gtgctttaat attttcagga
1981 gctgttattc tgttagaatt tataccagag attgcaatac ctgtattagg tacttttgca 2041 cttgtatcat atattgcgaa
taaggttcta accgttcaaa caatagataa tgctttaagt 2101 aaaagaaatg aaaaatggga tgaggtctat aaatatatag
taacaaattg gttagcaaag 2161 gttaatacac agattgatct aataagaaaa aaaatgaaag aagctttaga aaatcaagca
2221 gaagcaacaa aggctataat aaactatcag tataatcaat atactgagga agagaaaaat 2281 aatattaatt ttaatattga
tgatttaagt tcgaaactta atgagtctat aaataaagct 2341 atgattaata taaataaatt tttgaatcaa tgctctgttt catatttaat
gaattctatg 2401 atcccttatg gtgttaaacg gttagaagat tttgatgcta gtcttaaaga tgcattatta 2461 aagtatatat
atgataatag aggaacttta attggtcaag tagatagatt aaaagataaa 2521 gttaataata cacttagtac agatatacct tttcagcttt
ccaaatacgt agataatcaa 2581 agattattat ctacatttac tgaatatatt aagtaa
http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?db=nucleotide&val=18251975
Amino Acid Sequence
MQFVNKQFNYKDPVNGVDIAYIKIPNAGQMQPVKAFKIHN
KIWVIPERDTFTNPEEGDLNPPPEAKQVPVSYYDSTYLST
DNEKDNYLKGVTKLFERIYSTDLGRMLLTSIVRGIPFWGG
STIDTELKVIDTNCINVIQPDGSYRSEELNLVIIGPSADIIQFE
CKSFGHEVLNLTRNGYGSTQYIRFSPDFTFGFEESLEVDT
NPLLGAGKFATDPAVTLAHELIHAGHRLYGIAINPNRVFKV
NTNAYYEMSGLEVSFEELRTFGGHDAKFIDSLQENEFRLY
YYNKFKDIASTLNKAKSIVGTTASLQYMKNVFKEKYLLSE
DTSGKFSVDKLKFDKLYKMLTEIYTEDNFVKFFKVLNRKT
YLNFDKAVFKINIVPKVNYTIYDGFNLRNTNLAANFNGQNT
EINNMNFTKLKNFTGLFEFYKLLCVRGIITSKTKSLDKGYN
KALNDLCIKVNNWDLFFSPSEDNFTNDLNKGEEITSDTNIE
AAEENISLDLIQQYYLTFNFDNEPENISIENLSSDIIGQLELM
PNIERFPNGKKYELDKYTMFHYLRAQEFEHGKSRIALTNS
VNEALLNPSRVYTFFSSDYVKKVNKATEAAMFLGWVEQL
VYDFTDETSEVSTTDKIADITIIIPYIGPALNIGNMLYKDDFV
GALIFSGAVILLEFIPEIAIPVLGTFALVSYIANKVLTVQTIDN
ALSKRNEKWDEVYKYIVTNWLAKVNTQIDLIRKKMKEALE
NQAEATKAIINYQYNQYTEEEKNNINFNIDDLSSKLNESIN
KAMININKFLNQCSVSYLMNSMIPYGVKRLEDFDASLKDA
LLKYIYDNRGTLIGQVDRLKDKVNNTLSTDIPFQLSKY
VDNQRLLSTFTEYIK
Examine Properties
 Check for regions with like
charges or polarity
 Try to locate sulfur-sulfur and
salt bridges
 Look for possible a-helices, bsheets
Examine Properties
MQFVNKQFNYKDPVNGVDIAYIKIPNAGQMQPVKAFKIHNK
IWVIPERDTFTNPEEGDLNPPPEAKQVPVSYYDSTYLSTDN
EKDNYLKGVTKLFERIYSTDLGRMLLTSIVRGIPFWGGSTID
TELKVIDTNCINVIQPDGSYRSEELNLVIIGPSADIIQFECKSF
GHEVLNLTRNGYGSTQYIRFSPDFTFGFEESLEVDTNPLLG
AGKFATDPAVTLAHELIHAGHRLYGIAINPNRVFKVNTNAYY
EMSGLEVSFEELRTFGGHDAKFIDSLQENEFRLYYYNKFK
DIASTLNKAKSIVGTTASLQYMKNVFKEKYLLSEDTSGKFS
VDKLKFDKLYKMLTEIYTEDNFVKFFKVLNRKTYLNFDKAV
FKINIVPKVNYTIYDGFNLRNTNLAANFNGQNTEINNMNFTK
LKNFTGLFEFYKLLCVRGIITSKTKSLDKGYNKALNDLCIKV
NNWDLFFSPSEDNFTNDLNKGEEITSDTNIEAAEENISLDLI
QQYYLTFNFDNEPENISIENLSSDIIGQLELMPNIERFPNGK
KYELDKYTMFHYLRAQEFEHGKSRIALTNSVNEALLNPSR
VYTFFSSDYVKKVNKATEAAMFLGWVEQLVYDFTDETSEV
STTDKIADITIIIPYIGPALNIGNMLYKDDFVGALIFSGAVILLE
FIPEIAIPVLGTFALVSYIANKVLTVQTIDNALSKRNEKWDEV
YKYIVTNWLAKVNTQIDLIRKKMKEALENQAEATKAIINYQY
NQYTEEEKNNINFNIDDLSSKLNESINKAMININKFLNQCSV
SYLMNSMIPYGVKRLEDFDASLKDALLKYIYDNRGTLIGQV
DRLKDKVNNTLSTDIPFQLSKY VDNQRLLSTFTEYIK
Check for Homology
 Does your novel protein have
and close relatives?
 Similar proteins often have
similar structures and functions
 Input a.a. sequence into BLAST to
find matches
Check for Homology

BLAST http://www.ncbi.nlm.nih.gov/BLAST
Construct Model
 Capture images of the protein
 X-ray crystallography
 Pure protein crystal is exposed to xray, resulting diffraction patterns form
spots corresponding with e- density
 NMR Spectroscopy
 Sample is immersed in magnetic field,
causing nuclei to spin; variations in
spin denote different connectivity of
atoms
Construct Model
 Mathematical algorithms
 Software packages can calculate
thermodynamically stable
conformations of molecules
 Generates Cartesian coordinates of
atoms and outputs them as file
 Common molecular graphics
programs include
 Rasmol, Rastop and Cn3D
 Takes coordinate files and generates
image
Images
 RasMol viewer and ASN files becoming
standard, Cn3D growing in popularity
 2D *.png adapted from 3D binary *.asn
Chen J, Anderson JB, DeWeese-Scott C, Fedorova ND, Geer LY, He S, Hurwitz DI, Jackson JD, Jacobs AR, Lanczycki CJ, Liebert CA, Liu C, Madej T, Marchler-Bauer A, Marchler GH,
Mazumder R, Nikolskaya AN, Rao BS, Panchenko AR, Shoemaker BA, Simonyan V, Song JS, Thiessen PA, Vasudevan S, Wang Y, Yamashita RA, Yin JJ, Bryant SH, "MMDB: Entrez's
3D-structure database", Nucleic Acids Res. 2003 Jan; 31(1): 474-7.
Protein Data Bank
 Online repository of 3D protein
structures www.rcsb.org/pdb
 Search for related proteins and
structures
 Upload structure of your novel
protein with PDB information
Generate Hypotheses
 What do proteins similar to this
do?
 What purposes can large
patches of like residues serve?
 Charged patches can direct ion
flow
 Multiple sulfur and salt bridges
strengthen protein, resist high
temp.
Generate Hypotheses
 Our novel sequence is similar to:
bontoxilysin; botulinum neurotoxin
serotypes A, B & E, HHV7 alkaline
exonuclease and Zn finger protein 577
 Some are known neurotoxins
 Exonucleases cleave DNA
 Zinc fingers bind DNA
 So what might our novel protein do?
 Most search hits suggest neurotoxin
Our “novel” gene was none other than…
Botulinum toxin type A
(aka Botox!)