6. Run multiple alignment programs using the attached glutathione peroxidases (gluperox).

Start with Clustal Omega.

a. Which sequences appear to have a longer N-terminus than the others?

b. An asterisk represents a conserved position that is identical in all sequences. How many

such positions are present?

c. Compare the Clustal Omega output to the gluperox image. The researchers used ClustalW

to produce that figure. How close is the output to what is seen in the figure? Can you find any

major differences (other than order of sequences)?

d. The sequences all have one selenocysteine residue (u). At what amino acid

position is selenocysteine (use gi number 285803075 as your reference

sequence to determine the position)?

e. Is selenocysteine aligned perfectly by Clustal?

f. Name the amino acid that comes immediately before selenocysteine in most sequences.

Then name the amino acid that follows selenocysteine in most sequences.

g. Now run Muscle. Use the file gluoperox_2 because MUSCLE does not work with

selenocycsteine (the U is replaced by X). Comment on how different the alignment looks to the

Clustal alignment.

h. The X represents selenocysteine. Does the X align perfectly?

i. Now run T-Coffee (http://tcoffee.crg.cat/apps/tcoffee/do:regular ). Is the TCoffee output

closer to Muscle or Clustal?

j. Is the selenocysteine perfectly aligned? If not, where does it differ (what sequences)?

Reference

Gluperox

>gi|285803075|gb|ADC35417.1| glutathione peroxidase [Pinctada fucata] MASGTKSQKIISPVKIENFHQFSAKTLQGETINFKQYEGKVILVENTASLUGVTEREMRAMNELVDKFDG RLVILGFPCNQFGKQENCTNKEILNCLKYVRPGDGYEPKFPLFEKRDVNGEKADPIFKFLRDRLPIPCDE STSLMTDPKSIIWSPVTRSDISWNFEKFLIDPTGKPHRRFSRYSQTKGLEKDVKTLIDEFNVV >gi|264670920|gb|ACY72387.1| selenium-dependent glutathione peroxidase [Hyriopsis cumingii] MAVRIPLKPQNFFDVTAKTLKGQEVKLSKYKGKVILVENVASLUGTTVRDFTQMNELASKFEGKLVILGF PCNQFGHQENGNNDEILNSLKYVRPGNGFEPAFPIMEKCDVNGSNSHPLFQFLREKLQQPSDDPVSLMTH ANNIIWEPVTRTDISWNFEKFLIAPDGKPYKRYSRKFETKDIYNDIKDLIQQFGL >gi|111572533|gb|ABH10623.1| selenium-dependent glutathione peroxidase [Unio tumidus] MAWEPSQAALALVAFCGVLYFAAGEEGRTKCTNTKNHTVHDFSFLNVYGNETIDLRYYRGEVLLVVNVAT YUGLTVQYHGSNALQGKYRNDSFRVLGVPCSQFHFQEPAFTSEELMNGLKYARPGHGFVPNFNLTQKTEI NGHKEHPLYTYIKSECPPARDRFVQPILYEPIYTSDVRWNFEKFLIGRDGHPVYRYASTIDPRTSQMLDA DIAVEIKKTLHGHGKDIVDIVG >gi|145559053|gb|ABP73388.1| selenium-dependent glutathione peroxidase [Dreissena polymorpha] MRGDGVLSALVSLAAIVGSTVGSEADNAIPPRQACRPPLGNQTIFDFSIRNVYDNATIDMSQFRGKLTLV VNVATYUGLVVQYHGLNALQTQHGADGFQVLGVPCNQFHYQEPAWNGVELTNGLKHMRPGGGFVPNFPLT LTTKVNGQDEHPMYKYLKSLCKSVYKKLYHPILYDPVYTEDVKWNYEKFLIGPDGIPIYRYAQTVDPATD AQFLADVKEELAKLKGHTHGPSSAPSAHVPVVG >gi|222875570|gb|ACM68948.1| selenium-dependent glutathione peroxidase [Macrobrachium rosenbergii] MSAIKSFYELSAKALSGETVSFSKYKGKVILVENTAALUGTTTRDFVQMNELKEKFGDALAVLAFPCNQF GHQENNTDEELLNTLKYVRPGNNFVPKMEVFGKVTVNGSEAHPIFKYLKERLPLPSDDSVSFMNDPQCII WKPVCRTDIAWNFEKFLIGKDGQPFKRYSKKYETKNIEEDINALLK >gi|259124047|gb|ACV93251.1| glutathione peroxidase [Oreochromis niloticus] MAGHLKRFYDLTAKLLSGETLRFSALKDKVVLIENVASLUGTTTRDYTQVNDLHSRYSAEGLVILGVPCN QFGHQENCKNDEILRSLKYVRPGNGFEPKFQLLEKVDVNGKDAHPLFVYLKEKLPFPCDDALGLMNDPKY IIWSPVCRNDVSWNFEKFLVGPDGEPYKRYSRNFLTSDIEADIKQLLKRIK >gi|32363469|gb|AAP80181.1| glutathione peroxidase 1 [Homo sapiens] MCAARLAAAAAAAQSVYAFSARPLAGGEPVSLGSLRGKVLLIENVASLUGTTVRDYTQMNELQRRLGPRG LVVLGFPCNQFGHQENAKNEEILNSLKYVRPGGGFEPNFMLFEKCEVNGAGAHPLFAFLREALPAPSDDA TALMTDPKLITWSPVCRNDVAWNFEKFLVGPDGVPLRRYSRRFQTIDIEPDIEALLSQGPSCA >gi|169403976|ref|NP_001007282.2| glutathione peroxidase 1 [Danio rerio] MAGTMKKFYDLSAKLLSGDLLNFSSLKGKVVLIENVASLUGTTVRDYTQMNELHSRYADQGLVVLGAPCN QFGHQENCKNEEILQSLKYVRPGNGFEPKFQILEKLEVNGENAHPLFAFLKEKLPQPSDDPVSLMGDPKF IIWSPVCRNDISWNFEKFLIGPDGEPFKRYSRRFLTIDIDADIKELLKRTK >gi|84871986|ref|NP_032186.2| glutathione peroxidase 1 [Mus musculus] MCAARLSAAAQSTVYAFSARPLTGGEPVSLGSLRGKVLLIENVASLUGTTIRDYTEMNDLQKRLGPRGLV VLGFPCNQFGHQENGKNEEILNSLKYVRPGGGFEPNFTLFEKCEVNGEKAHPLFTFLRNALPTPSDDPTA LMTDPKYIIWSPVCRNDIAWNFEKFLVGPDGVPVRRYSRRFRTIDIEPDIETLLSQQSGNS >gi|32967607|ref|NP_002074.2| glutathione peroxidase 2 [Homo sapiens] MAFIAKSFYDLSAISLDGEKVDFNTFRGRAVLIENVASLUGTTTRDFTQLNELQCRFPRRLVVLGFPCNQ FGHQENCQNEEILNSLKYVRPGGGYQPTFTLVQKCEVNGQNEHPVFAYLKDKLPYPYDDPFSLMTDPKLI IWSPVRRSDVAWNFEKFLIGPEGEPFRRYSRTFPTINIEPDIKRLLKVAI >gi|34783711|gb|AAH54848.2| Glutathione peroxidase 2 [Mus musculus] MAYIAKSFYDLSAIGLDGEKIDFNTFRGRAVLIENVASLUGTTTRDYNQLNELQCRFPRRLVVLGFPCNQ FGHQENCQNEEILNSLKYVRPGGGYQPTFSLTQKCDVNGQNEHPVFAYLKDKLPYPYDDPFSLMTDPKLI IWSPVRRSDVSWNFEKFLIGPEGEPFRRYSRSFQTINIEPDIKRLLKVAI >gi|312600934|gb|ADQ92353.1| glutathione peroxidase [Mizuhopecten yessoensis] MATVVSVRSPVKLQNFFDFHAKTLDGETINFSKYKGKVILVMNVASLUDTTVRDYTQMNELVQKYGDKLV VLGFPCNQFGHQENTKNHEILTALKYLRPGDGFVPKFPIFEKLEVNGKNASEIFVFLRNHLPLPSDDPTS FMKSCSSIIWEPVTRTDLAWNFEKFLITPEGKPYKRFSRYYLTCNLQSEIKTLIEKFNVK

Gluperox 2

>gi|285803075|gb|ADC35417.1| glutathione peroxidase [Pinctada fucata] MASGTKSQKIISPVKIENFHQFSAKTLQGETINFKQYEGKVILVENTASLXGVTEREMRAMNELVDKFDG RLVILGFPCNQFGKQENCTNKEILNCLKYVRPGDGYEPKFPLFEKRDVNGEKADPIFKFLRDRLPIPCDE STSLMTDPKSIIWSPVTRSDISWNFEKFLIDPTGKPHRRFSRYSQTKGLEKDVKTLIDEFNVV >gi|264670920|gb|ACY72387.1| selenium-dependent glutathione peroxidase [Hyriopsis cumingii] MAVRIPLKPQNFFDVTAKTLKGQEVKLSKYKGKVILVENVASLXGTTVRDFTQMNELASKFEGKLVILGF PCNQFGHQENGNNDEILNSLKYVRPGNGFEPAFPIMEKCDVNGSNSHPLFQFLREKLQQPSDDPVSLMTH ANNIIWEPVTRTDISWNFEKFLIAPDGKPYKRYSRKFETKDIYNDIKDLIQQFGL >gi|111572533|gb|ABH10623.1| selenium-dependent glutathione peroxidase [Unio tumidus] MAWEPSQAALALVAFCGVLYFAAGEEGRTKCTNTKNHTVHDFSFLNVYGNETIDLRYYRGEVLLVVNVAT YXGLTVQYHGSNALQGKYRNDSFRVLGVPCSQFHFQEPAFTSEELMNGLKYARPGHGFVPNFNLTQKTEI NGHKEHPLYTYIKSECPPARDRFVQPILYEPIYTSDVRWNFEKFLIGRDGHPVYRYASTIDPRTSQMLDA DIAVEIKKTLHGHGKDIVDIVG >gi|145559053|gb|ABP73388.1| selenium-dependent glutathione peroxidase [Dreissena polymorpha] MRGDGVLSALVSLAAIVGSTVGSEADNAIPPRQACRPPLGNQTIFDFSIRNVYDNATIDMSQFRGKLTLV VNVATYXGLVVQYHGLNALQTQHGADGFQVLGVPCNQFHYQEPAWNGVELTNGLKHMRPGGGFVPNFPLT LTTKVNGQDEHPMYKYLKSLCKSVYKKLYHPILYDPVYTEDVKWNYEKFLIGPDGIPIYRYAQTVDPATD AQFLADVKEELAKLKGHTHGPSSAPSAHVPVVG >gi|222875570|gb|ACM68948.1| selenium-dependent glutathione peroxidase [Macrobrachium rosenbergii] MSAIKSFYELSAKALSGETVSFSKYKGKVILVENTAALXGTTTRDFVQMNELKEKFGDALAVLAFPCNQF GHQENNTDEELLNTLKYVRPGNNFVPKMEVFGKVTVNGSEAHPIFKYLKERLPLPSDDSVSFMNDPQCII WKPVCRTDIAWNFEKFLIGKDGQPFKRYSKKYETKNIEEDINALLK >gi|259124047|gb|ACV93251.1| glutathione peroxidase [Oreochromis niloticus] MAGHLKRFYDLTAKLLSGETLRFSALKDKVVLIENVASLXGTTTRDYTQVNDLHSRYSAEGLVILGVPCN QFGHQENCKNDEILRSLKYVRPGNGFEPKFQLLEKVDVNGKDAHPLFVYLKEKLPFPCDDALGLMNDPKY IIWSPVCRNDVSWNFEKFLVGPDGEPYKRYSRNFLTSDIEADIKQLLKRIK >gi|32363469|gb|AAP80181.1| glutathione peroxidase 1 [Homo sapiens] MCAARLAAAAAAAQSVYAFSARPLAGGEPVSLGSLRGKVLLIENVASLXGTTVRDYTQMNELQRRLGPRG LVVLGFPCNQFGHQENAKNEEILNSLKYVRPGGGFEPNFMLFEKCEVNGAGAHPLFAFLREALPAPSDDA TALMTDPKLITWSPVCRNDVAWNFEKFLVGPDGVPLRRYSRRFQTIDIEPDIEALLSQGPSCA >gi|169403976|ref|NP_001007282.2| glutathione peroxidase 1 [Danio rerio] MAGTMKKFYDLSAKLLSGDLLNFSSLKGKVVLIENVASLXGTTVRDYTQMNELHSRYADQGLVVLGAPCN QFGHQENCKNEEILQSLKYVRPGNGFEPKFQILEKLEVNGENAHPLFAFLKEKLPQPSDDPVSLMGDPKF IIWSPVCRNDISWNFEKFLIGPDGEPFKRYSRRFLTIDIDADIKELLKRTK >gi|84871986|ref|NP_032186.2| glutathione peroxidase 1 [Mus musculus] MCAARLSAAAQSTVYAFSARPLTGGEPVSLGSLRGKVLLIENVASLXGTTIRDYTEMNDLQKRLGPRGLV VLGFPCNQFGHQENGKNEEILNSLKYVRPGGGFEPNFTLFEKCEVNGEKAHPLFTFLRNALPTPSDDPTA LMTDPKYIIWSPVCRNDIAWNFEKFLVGPDGVPVRRYSRRFRTIDIEPDIETLLSQQSGNS >gi|32967607|ref|NP_002074.2| glutathione peroxidase 2 [Homo sapiens] MAFIAKSFYDLSAISLDGEKVDFNTFRGRAVLIENVASLXGTTTRDFTQLNELQCRFPRRLVVLGFPCNQ FGHQENCQNEEILNSLKYVRPGGGYQPTFTLVQKCEVNGQNEHPVFAYLKDKLPYPYDDPFSLMTDPKLI IWSPVRRSDVAWNFEKFLIGPEGEPFRRYSRTFPTINIEPDIKRLLKVAI >gi|34783711|gb|AAH54848.2| Glutathione peroxidase 2 [Mus musculus] MAYIAKSFYDLSAIGLDGEKIDFNTFRGRAVLIENVASLXGTTTRDYNQLNELQCRFPRRLVVLGFPCNQ FGHQENCQNEEILNSLKYVRPGGGYQPTFSLTQKCDVNGQNEHPVFAYLKDKLPYPYDDPFSLMTDPKLI IWSPVRRSDVSWNFEKFLIGPEGEPFRRYSRSFQTINIEPDIKRLLKVAI >gi|312600934|gb|ADQ92353.1| glutathione peroxidase [Mizuhopecten yessoensis] MATVVSVRSPVKLQNFFDFHAKTLDGETINFSKYKGKVILVMNVASLXDTTVRDYTQMNELVQKYGDKLV VLGFPCNQFGHQENTKNHEILTALKYLRPGDGFVPKFPIFEKLEVNGKNASEIFVFLRNHLPLPSDDPTS FMKSCSSIIWEPVTRTDLAWNFEKFLITPEGKPYKRFSRYYLTCNLQSEIKTLIEKFNVK

Image