Example from Kibak
Example Integrative Assignment Part II Using Cytochrome b Instead of c
Figures 1 through 4 are binary alignments of Cytochrome b from the example organism I chose,
Strongylocentrotus purpuratus, with representative Cytochromes b from four major phylogenetic groups.
Figure 5 is a multiple alignment of all five sequences.
Strongylocentrotus
Homo
MAAPLRKEHPIFRILNSTFVDLPLPSNLSIWWNSGSLLGLCLVVQILTGIFLAMHYTADI
M-TPMRKTNPLMKLINHSFIDLPTPSNISAWWNFGSLLGACLILQITTGLFLAMHYSPDA
* ** *
* * *** *** * *** ***** ** ** ** ****** *
Strongylocentrotus
Homo
TLAFSSVMHILRDVNYGWFLRYVHANGVSLFFICMYCHIGRGLYYGSYNKIETWNVGVIL
STAFSSIAHITRDVNYGWIIRYLHANGASMFFICLFLHIGRGLYYGSFLYSETWNIGIIL
**** ** ******* ** **** * ****
**********
**** * **
Strongylocentrotus
Homo
FLVTILTAFMGYVLVWGQMSFWAATVITNLVSAIPYIGTIIVQWLWGGFSVDNATLTRFF
LLATMATAFMGYVLPWGQMSFWGATVITNLLSAIPYIGTDLVQWIWGGYSVDSPTLTRFF
* * ******** ******* ******* ******** *** *** *** ******
Strongylocentrotus
Homo
PFHFLFPFIIAALAVIHLVFLHNSGANNPFAFNSNYDKAPFHIYFTTKDTVGFILLVAAL
TFHFILPFIIAALATLHLLFLHETGSNNPLGITSHSDKITFHPYYTIKDALGLLLFLLSL
*** ******** ** *** * ***
* ** ** * * ** * *
*
Strongylocentrotus
Homo
FSLALLFPGALNDPENFIPANPLVTPPHIQPEWYFLFAYAILRSIPNKLGGVIALVAAIL
MTLTLFSPDLLGDPDNYTLANPLNTPPHIKPEWYFLFAYTILRSVPNKLGGVLALLLSIL
* * * * ** *
**** ***** ********* **** ******* **
**
Strongylocentrotus
Homo
VLFLMPLLNTSKNESNSFRPLSQAAFWLLVAHLFILTWIGSQPVEYPYVLLGQVASVLYF
ILAMIPILHMSKQQSMMFRPLSQSLYWLLAADLLILTWIGGQPVSYPFTIIGQVASVLYF
*
* * ** * ******
*** * * ****** *** **
*********
Strongylocentrotus
Homo
SLFIFGFPIVSSIENKIIFS
TTILILMPTISLIENKMLK* * ****
Figure 1. Cytochrome b from Strongylocentrotus purpuratus compared to the human sequence. An “*” means the sequence is
identical at that position. There were 237 exact matches out of 380 (62.4% identical). The longest stretch of identical amino acids
(10) is shaded.
The other alignments continue on the next page.
Example from Kibak
Strongylocentrotus
Arabidopsis
MAAPLR----KEHPIFRILNSTFVDLPLPSNLSIWWNSGSLLGLCLVVQILTGIFLAMHY
MTIRNQRFSLLKQPISSTLNQHLVDYPTPSNLSYWWGFGPLAGICLVIQIVTGVFLAMHY
*
**
**
** * ***** ** * * * *** ** ** ******
Strongylocentrotus
Arabidopsis
TADITLAFSSVMHILRDVNYGWFLRYVHANGVSLFFICMYCHIGRGLYYGSYN--KIETW
TPHVDLAFNSVEHIMRDVEGGWLLRYMHANGASMFFIVVYLHIFRGLYYASYSSPREFVW
*
*** ** ** *** ** *** **** * *** * ** ***** **
*
Strongylocentrotus
Arabidopsis
NVGVILFLVTILTAFMGYVLVWGQMSFWAATVITNLVSAIPYIGTIIVQWLWGGFSVDNA
CLGVVIFLLMIVTAFIGYVLPWGQMSFWGATVITSLASAIPVVGDTIVTWLWGGFSVDNA
** ** * *** **** ******* ***** * **** * ** ***********
Strongylocentrotus
Arabidopsis
TLTRFFPFHFLFPFIIAALAVIHLVFLHNSGANNPFAFNSNYDKAPFHIYFTTKDTVGFI
TLNRFFSLHYLLPFILVGASLLHLAALHQYGSNNPLGVHSEMDKIAFYPYFYVKDLVGWV
** *** * * ***
** ** * ***
* ** * ** ** **
Strongylocentrotus
Arabidopsis
LLVAALFSLALLFPGALNDPENFIPANPLVTPPHIQPEWYFLFAYAILRSIPNKLGGVIA
AFAIFFSIWIFYAPNVLGHPDNYIPANPMSTPPHIVPEWYFLPIYAILRSIPDKAGGVAA
* * * * ***** ***** ****** ******** * *** *
Strongylocentrotus
Arabidopsis
LVAAILVLFLMPLLNTSKNESNSFRPLSQAAFWLLVAHLFILTWIGSQPVEYPYVLLGQV
IALVFICLLALPFFKSMYVRSSSFRPIYQGMFWLLLADCLLLGWIGCQPVEAPFVTIGQI
*
*
* **** * **** *
* *** **** * * **
Strongylocentrotus
Arabidopsis
ASVLYFSLFIFGFPIVSSIENKIIFS-------SSLVFFLFFAI-TPILGRVGRGIPNSYTDETDHT
*
* *
**
* *
Figure 2. Cytochrome b from Strongylocentrotus purpuratus compared to the Arabidopsis sequence. There were 202 exact
matches out of the 380 Strongylocentrotus positions (53.2% identical). The longest stretch of identical amino acids (13) is
shaded.
Strongylocentrotus
Saccharomyces
MAAPLRKEHPIFRILNSTFVDLPLPSNLSIWWNSGSLLGLCLVVQILTGIFLAMHYTADI
--MAFRKSNVYLSLVNSYVIDSPQPSSINYWWNMGSLLGLCLVIQIVTGIFMAMHYSSNI
**
**
* * **
*** ********* ** **** ****
*
Strongylocentrotus
Saccharomyces
TLAFSSVMHILRDVNYGWFLRYVHANGVSLFFICMYCHIGRGLYYGSYN--KIETWNVGV
ELAFSSVEHIMRDVHNGYILRYLHANGASFFFMVMFMHMAKGLYYGSYRSPRVTLWNVGV
****** ** *** * *** **** * ** * *
*******
*****
Strongylocentrotus
Saccharomyces
ILFLVTILTAFMGYVLVWGQMSFWAATVITNLVSAIPYIGTIIVQWLWGGFSVDNATLTR
IIFILTIATAFLGYCCVYGQMSHWGATVITNLFSAIPFVGNDIVSWLWGGFSVSNPTIQR
* * ** *** ** * **** * ******* **** * ** ******** * * *
Strongylocentrotus
Saccharomyces
FFPFHFLFPFIIAALAVIHLVFLHNSGANNPFAFNSNYDKAPFHIYFTTKDTVGFILLVA
FFALHYLVPFIIAAMVIMHLMALHIHGSSNPLGITGNLDRIPMHSYFIFKDLVTVFLFML
** * * ******
** ** * **
* * * * ** ** *
*
Strongylocentrotus
Saccharomyces
ALFSLALLFPGALNDPENFIPANPLVTPPHIQPEWYFLFAYAILRSIPNKLGGVIALVAA
ILALFVFYSPNTLGHPDNYIPGNPLVTPASIVPEWYLLPFYAILRSIPDKLLGVITMFAA
*
* * * * ** ****** * **** * ******** ** ***
**
Strongylocentrotus
Saccharomyces
ILVLFLMPLLNTSKNESNSFRPLSQAAFWLLVAHLFILTWIGSQPVEYPYVLLGQVASVL
ILVLLVLPFTDRSVVRGNTFKVLSKFFFFIFVFNFVLLGQIGACHVEVPYVLMGQIATFI
****
*
*
* * **
*
*
* **
** **** ** *
Strongylocentrotus
Saccharomyces
YFSLFIFGFPIVSSIENKIIFS----YFAYFLIIVPVISTIENVLFYIGRVNK
** *
* * ***
Figure 3. Cytochrome b from Strongylocentrotus purpuratus compared to the Baker’s Yeast sequence. There were 200 exact
matches out of the 380 Strongylocentrotus positions (52.6% identical). The longest stretch of identical amino acids (9) is shaded.
Example from Kibak
Strongylocentrotus
Apis
--MAAPLRKEHPIFRILNSTFVDLPLPSNLSIWWNSGSLLGLCLVVQILTGIFLAMHYTA
MKKFMNFFSSNEFLKMIMST-IYLPTPVNINYMWNFGSILGIFLMIQIISGFILSMHYCP
**
** * *
** ** ** * ** * * ***
Strongylocentrotus
Apis
DITLAFSSVMHILRDVNYGWFLRYVHANGVSLFFICMYCHIGRGLYYGSYNKIETWNVGV
NIDIAFWSITNIMKDMNSGWLFRLIHMNGASFYFLMMYIHISRNLFYCSYKLNNVWGIGI
* ** *
* * * ** * * ** * * ** ** * * * **
* *
Strongylocentrotus
Apis
ILFLVTILTAFMGYVLVWGQMSFWAATVITNLVSAIPYIGTIIVQWLWGGFSVDNATLTR
MILLMSMAAAFMGYVLPWGQMSYWGATVITNLLSAIPYIGDTIVLWIWGGFSINNATLNR
*
******* ***** * ******* ******* ** * ***** **** *
Strongylocentrotus
Apis
FFPFHFLFPFIIAALAVIHLVFLHNSGANNPFAFNSNYDKAPFHIYFTTKDTVGFILLVA
FFSLHFILPLLILFMVILHLFALHLTGSSNPLGSNFNNYKISFHPYFSIKDLLGFYIILF
** ** * *
** ** * **
* * * ** ** ** **
Strongylocentrotus
Apis
ALFSLALLFPGALNDPENFIPANPLVTPPHIQPEWYFLFAYAILRSIPNKLGGVIALVAA
IFMFINFQFPYHLGDPDNFKIANPMNTPTHIKPEWYFLFAYSILRAIPNKLGGVIGLVMS
** * ** ** *** ** ** ********* *** ********* **
Strongylocentrotus
Apis
ILVLFLMPLLNTSKNESNSFRPLSQAAFWLLVAHLFILTWIGSQPVEYPYVLLGQVASVL
ILILYIMIFYNN-KMMNNKFNMLNKIYYWMFINNFILLTWLGKQLIEYPFTNINMLFTTT
** * *
* *
* * *
*
*** * * ***
Strongylocentrotus
Apis
YFSLFIFGFPIVSSIENKIIFS--YFLYFFLNFYLSKLWDNLIWNSPLN
** *
*
* * *
Figure 4. Cytochrome b from Strongylocentrotus purpuratus compared to the Honey Bee sequence. There were 173 exact
matches out of the 380 Strongylocentrotus positions (45.5% identical). The two longest stretches of identical amino acids (9) are
shaded.
Based on these four two-way alignments and the percent identity between the sea urchin sequence and the
others, the Purple Sea Urchin Cytochrome b is most similar to the Human Cytochrome b. This is not
surprising as both of these species are deuterostomes and would be considered the most closely related of
these species by traditional classification schemes. What is surprising is that the most dissimilar sequence
based on these alignments is the Honey Bee, also a metazoan, while the Yeast (fungi) and the Arabidopsis
(plant) are more similar than a fellow animal!
In order to identify the regions in the protein that remain unchanged over billions of years, Figure 5 on the
next page presents a multiple alignment of Cytochrome b from five very different organisms. There is
most variability at each end of the protein. Near the middle of the sequence there is a stretch of 19
positions where only four positions have been able to change since these five species diverged from their
common ancestor.
Here goes the section where you speculate on why those positions cannot change… I am not going to do
that for you! Think about it!
Example from Kibak
Strongylocentrotus
Homo
Arabidopsis
Saccharomyces
Apis
--MAAPLRKE------HPIFRILNSTFVDLPLPSNLSIWWNSGSLLGLCLVVQILTGIFL
---MTPMRKT------NPLMKLINHSFIDLPTPSNISAWWNFGSLLGACLILQITTGLFL
----MTIRNQRFSLLKQPISSTLNQHLVDYPTPSNLSYWWGFGPLAGICLVIQIVTGVFL
----MAFRKS------NVYLSLVNSYVIDSPQPSSINYWWNMGSLLGLCLVIQIVTGIFM
MKKFMNFFSS------NEFLKMIMSTIY-LPTPVNINYMWNFGSILGIFLMIQIISGFIL
* *
* *
* * ** *
Strongylocentrotus
Homo
Arabidopsis
Saccharomyces
Apis
AMHYTADITLAFSSVMHILRDVNYGWFLRYVHANGVSLFFICMYCHIGRGLYYGSYN--K
AMHYSPDASTAFSSIAHITRDVNYGWIIRYLHANGASMFFICLFLHIGRGLYYGSFL--Y
AMHYTPHVDLAFNSVEHIMRDVEGGWLLRYMHANGASMFFIVVYLHIFRGLYYASYSSPR
AMHYSSNIELAFSSVEHIMRDVHNGYILRYLHANGASFFFMVMFMHMAKGLYYGSYRSPR
SMHYCPNIDIAFWSITNIMKDMNSGWLFRLIHMNGASFYFLMMYIHISRNLFYCSYK--L
***
** *
* *
*
* * ** * *
*
* * *
Strongylocentrotus
Homo
Arabidopsis
Saccharomyces
Apis
IETWNVGVILFLVTILTAFMGYVLVWGQMSFWAATVITNLVSAIPYIGTIIVQWLWGGFS
SETWNIGIILLLATMATAFMGYVLPWGQMSFWGATVITNLLSAIPYIGTDLVQWIWGGYS
EFVWCLGVVIFLLMIVTAFIGYVLPWGQMSFWGATVITSLASAIPVVGDTIVTWLWGGFS
VTLWNVGVIIFILTIATAFLGYCCVYGQMSHWGATVITNLFSAIPFVGNDIVSWLWGGFS
NNVWGIGIMILLMSMAAAFMGYVLPWGQMSYWGATVITNLLSAIPYIGDTIVLWIWGGFS
* *
** **
**** * ***** * **** *
* * *** *
Strongylocentrotus
Homo
Arabidopsis
Saccharomyces
Apis
VDNATLTRFFPFHFLFPFIIAALAVIHLVFLHNSGANNPFAFNSNYDKAPFHIYFTTKDT
VDSPTLTRFFTFHFILPFIIAALATLHLLFLHETGSNNPLGITSHSDKITFHPYYTIKDA
VDNATLNRFFSLHYLLPFILVGASLLHLAALHQYGSNNPLGVHSEMDKIAFYPYFYVKDL
VSNPTIQRFFALHYLVPFIIAAMVIMHLMALHIHGSSNPLGITGNLDRIPMHSYFIFKDL
INNATLNRFFSLHFILPLLILFMVILHLFALHLTGSSNPLGSNFNNYKISFHPYFSIKDL
* *** *
*
** ** * **
*
**
Strongylocentrotus
Homo
Arabidopsis
Saccharomyces
Apis
VGFILLVAALFSLALLFPGALNDPENFIPANPLVTPPHIQPEWYFLFAYAILRSIPNKLG
LGLLLFLLSLMTLTLFSPDLLGDPDNYTLANPLNTPPHIKPEWYFLFAYTILRSVPNKLG
VGWVAFAIFFSIWIFYAPNVLGHPDNYIPANPMSTPPHIVPEWYFLPIYAILRSIPDKAG
VTVFLFMLILALFVFYSPNTLGHPDNYIPGNPLVTPASIVPEWYLLPFYAILRSIPDKLL
LGFYIILFIFMFINFQFPYHLGDPDNFKIANPMNTPTHIKPEWYFLFAYSILRAIPNKLG
* * * *
** ** * **** * * *** * *
Strongylocentrotus
Homo
Arabidopsis
Saccharomyces
Apis
GVIALVAAILVLFLMPLLNTSKNESNSFRPLSQAAFWLLVAHLFILTWIGSQPVEYPYVL
GVLALLLSILILAMIPILHMSKQQSMMFRPLSQSLYWLLAADLLILTWIGGQPVSYPFTI
GVAAIALVFICLLALPFFKSMYVRSSSFRPIYQGMFWLLLADCLLLGWIGCQPVEAPFVT
GVITMFAAILVLLVLPFTDRSVVRGNTFKVLSKFFFFIFVFNFVLLGQIGACHVEVPYVL
GVIGLVMSILILYIMIFYNN-KMMNNKFNMLNKIYYWMFINNFILLTWLGKQLIEYPFTN
**
*
*
*
*
*
Strongylocentrotus
Homo
Arabidopsis
Saccharomyces
Apis
LGQVASVLYFSLFIFGFPIVSSIENKIIFS-------IGQVASVLYFTTILILMPTISLIENKMLK--------IGQISSLVFFLFFAIT-PILGRVGRGIPNSYTDETDHT
MGQIATFIYFAYFLIIVPVISTIEN--VLFYIGRVNKINMLFTTTYFLYFFLNFYLSKLWDNLIWNSPLN----*
Figure 5. The Cytochrome b sequence of Purple Sea Urchin compared to the Cytochromes b of a deuterostome, a higher plant,
a fungus, and a protostome. The most similar region is shaded.