Difference between revisions of "Part:BBa K1080001:Design"

Revision as of 02:36, 24 September 2013

ChlH

Design Notes

CR-ChlH sequence with His Tag in pET15b. Sequence from Translation start site.

Note 8 PstI sites CTGCAG (785, 1031, 1385, 1652, 2681, 2825, 3458, 3569) No EcoRI or XbaI sites. 1 SpeI (ACTAGT) (4226) site 23 bp after stop codon.

ATGCGGGGTTCTCATCATCATCATCATCATGGTATGGCTAGCATGACTGGTGGACAGCAAATGGGTCGGGATCTGT

ACGACGATGACGATAAGGATCATCCCTTCACCGCGTGCAATGTGGCGACTGGACCCCGGCCGCCCATGACCACCTT

CACCGGTGGCAACAAGGGCCCTGCTAAGCAGCAGGTGTCGCTGGATCTGCGCGACGAGGGCGCTGGCATGTTCACC

AGCACCAGCCCGGAGATGCGCCGTGTCGTCCCTGACGATGTGAAGGGTCGCGTTAAGGTGAAGGTTGTGTACGTGG

TGCTGGAGGCCCAGTACCAGTCGGCCATCAGCGCTGCGGTGAAGAACATCAACGCCAAGAACTCCAAGGTGTGCTT

CGAGGTGGTGGGCTACCTGCTGGAGGAGCTGCGTGACCAGAAGAACCTCGATATGCTCAAGGAGGATGTGGCCTCT

GCCAACATCTTCATCGGCTCGCTCATCTTCATTGAGGAGCTTGCCGAGAAGATTGTGGAGGCGGTGAGCCCCCTGC

GCGAGAAGCTGGACGCGTGCCTGATCTTCCCGTCCATGCCGGCGGTCATGAAGCTGAACAAGCTGGGCACGTTTTC

GATGGCTCAGCTGGGCCAGTCGAAGTCGGTGTTCTCGGAGTTCATCAAGTCTGCTCGCAAGAACAACGACAACTTC

GAGGAGGGCTTGCTGAAGCTGGTGCGCACCCTGCCTAAGGTGCTGAAGTATCTGCCCTCGGACAAGGCGCAGGACG

CCAAGAACTTCGTGAACAGCCTGCAGTACTGGCTGGGCGGTAACTCGGACAACCTGGAGAACCTGCTGCTGAACAC

CGTCAGCAACTACGTGCCCGCTCTGAAGGGCGTGGACTTCAGCGTGGCTGAGCCCACCGCCTACCCCGATGTGGGT

ATCTGGCACCCTCTGGCCTCGGGCATGTACGAGGACCTGAAGGAGTACCTGAACTGGTACGACACCCGCAAGGACA

TGGTCTTCGCCAAGGACGCCCCCGTCATTGGCCTGGTGCTGCAGCGCTCGCACCTGGTGACTGGCGATGAGGGCCA

CTACAGCGGCGTGGTCGCTGAGCTGGAGAGCCGCGGTGCTAAGGTCATCCCCGTCTTTGCCGGTGGCCTGGACTTC

TCCGCCCCCGTCAAGAAGTTCTTCTACGACCCCCTGGGCTCTGGCCGCACGTTCGTGGACACCGTTGTGTCGCTGA

CCGGCTTCGCGCTGGTGGGCGGCCCCGCGCGCCAGGACGCGCCGAAGGCCATTGAGGCGCTGAAGAACCTGAACGT

GCCCTACCTGGTGTCGCTGCCGCTGGTGTTCCAGACCACTGAGGAGTGGCTGGACAGCGAGCTGGGCGTGCACCCC

GTCCAGGTGGCTCTGCAGGTTGCCCTGCCCGAGCTGGATGGTGCCATGGAGCCCATCGTGTTCGCTGGCCGTGACT

CGAACACCGGCAAGTCGCACTCGCTGCCCGACCGCATCGCTTCGCTGTGCGCTCGCGCCGTGAACTGGGCCAACCT

GCGCAAGAAGCGCAACGCCGAGAAGAAGCTGGCCGTCACCGTGTTCAGCTTCCCCCCTGACAAGGGCAACGTCGGC

ACTGCCGCCTACCTGAACGTGTTCGGCTCCATCTACCGCGTGCTGAAGAACCTGCAGCGCGAGGGCTACGACGTGG

GCGCCCTGCCGCCCTCGGAGGAGGATCTGATCCAGTCGGTGCTGACCCAGAAGGAGGCCAAGTTCAACTCGACCGA

CCTGCACATCGCCTACAAGATGAAGGTGGACGAGTACCAGAAGCTGTGCCCTTACGCCGAGGCGCTGGAGGAGAAC

TGGGGCAAGCCCCCCGGCACCCTGAACACCAACGGCCAGGAGCTGCTGGTGTACGGCCGCCAGTACGGCAACGTCT

TCATCGGCGTGCAGCCCACCTTCGGCTACGAGGGCGACCCGATGCGCCTGCTGTTCTCGAAGTCGGCCAGCCCCCA

CCACGGCTTCGCCGCCTACTACACCTTCCTGGAGAAGATCTTCAAGGCCGACGCCGTGCTGCACTTCGGCACCCAC

GGCTCGCTGGAGTTCATGCCCGGCAAGCAGGTCGGCATGTCGGGTGTGTGCTACCCCGACTCGCTGATCGGCACCA

TCCCCAACCTCTACTACTACGCCGCCAACAACCCGTCTGAGGCCACCATCGCCAAGCGCCGCTCGTACGCCAACAC

CATTTCGTACCTGACGCCGCCTGCCGAGAACGCCGGCCTGTACAAGGGCCTGAAGGAGCTGAAGGAGCTGATCAGC

TCGTACCAGGGCATGCGTGAGTCTGGCCGCGCCGAGCAGATCTGCGCCACCATCATTGAGACCGCCAAGCTGTGCA

ACCTGGACCGCGACGTGACCCTGCCCGACGCTGACGCCAAGGACCTGACCATGGACATGCGCGACAGCGTTGTGGG

CCAGGTGTACCGCAAGCTGATGGAGATTGAGTCCCGCCTGCTGCCCTGCGGCCTGCACGTGGTGGGCTGCCCGCCC

ACCGCCGAGGAGGCCGTGGCCACCCTGGTCAACATCGCTGAGCTGGACCGCCCGGACAACAACCCCCCCATCAAGG

GCATGCCCGGCATCCTGGCCCGCGCCATTGGTCGCGACATCGAGTCGATTTACAGCGGCAACAACAAGGGCGTCCT

GGCTGACGTTGACCAGCTGCAGCGCATCACCGAGGCCTCCCGCACCTGCGTGCGCGAGTTCGTGAAGGACCGCACC

GGCCTGAACGGCCGCATCGGCACCAACTGGATCACCAACCTGCTCAAGTTCACCGGCTTCTACGTGGACCCCTGGG

TGCGCGGCCTGCAGAACGGCGAGTTCGCCAGCGCCAACCGCGAGGAGCTGATCACCCTGTTCAACTACCTGGAGTT

CTGCCTGACCCAGGTGGTCAAGGACAACGAGCTGGGCGCCCTGGTAGAGGCGCTGAACGGCCAGTACGTCGAGCCC

GGCCCCGGCGGTGACCCCATCCGCAACCCCAACGTGCTGCCCACCGGCAAGAACATCCACGCCCTGGACCCTCAGT

CGATTCCCACTCAGGCCGCGCTGAAGAGCGCCCGCCTGGTGGTGGACCGCCTGCTGGACCGCGAGCGCGACAACAA

CGGCGGCAAGTACCCCGAGACCATCGCGCTGGTGCTGTGGGGCACTGACAACATCAAGACCTACGGCGAGTCGCTG

GCCCAGGTCATGATGATGGTCGGTGTCAAGCCCGTGGCCGACGCCCTGGGCCGCGTGAACAAGCTGGAGGTGATCC

CTCTGGAGGAGCTGGGCCGCCCCCGCGTGGACGTGGTTGTCAACTGCTCGGGTGTGTTCCGCGACCTGTTCGTGAA

CCAGATGCTGCTGCTGGACCGCGCCATCAAGCTGGCGGCCGAGCAGGACGAGCCCGATGAGATGAACTTCGTGCGC

AAGCACGCCAAGCAGCAGGCGGCGGAGCTGGGCCTGCAGAGCCTGCGCGACGCGGCCACCCGTGTGTTCTCCAACA

GCTCGGGCTCCTACTCGTCCAACGTCAACCTGGCGGTGGAGAACAGCAGCTGGAGCGACGAGTCGCAGCTGCAGGA

GATGTACCTGAAGCGCAAGTCGTACGCCTTCAACTCGGACCGCCCCGGCGCCGGTGGCGAGATGCAGCGCGACGTG

TTCGAGACGGCCATGAAGACCGTGGACGTGACCTTCCAGAACCTGGACTCGTCCGAGATCTCGCTGACCGATGTGT

CGCACTACTTCGACTCCGACCCCACCAAGCTGGTGGCGTCGCTGCGCAACGACGGCCGCACCCCCAACGCCTACAT

CGCCGACACCACCACCGCCAACGCGCAGGTCCGCACTCTGGGTGAGACCGTGCGCCTGGACGCCCGCACCAAGCTG

CTCAACCCCAAGTGGTACGAGGGCATGCTTGCCTCGGGCTACGAGGGCGTGCGCGAGATCCAGAAGCGCATGACCA

ACACCATGGGCTGGTCGGCCACCTCGGGCATGGTGGACAACTGGGTGTACGACGAGGCCAACTCGACCTTCATCGA

GGATGCGGCCATGGCCGAGCGCCTGATGAACACCAACCCCAACAGCTTCCGCAAGCTGGTGGCCACCTTCCTGGAG

GCCAACGGCCGCGGCTACTGGGACGCCAAGCCCGAGCAGCTGGAGCGCCTGCGCCAGCTGTACATGGACGTGGAGG

ACAAGATTGAGGGCGTCGAATAAGCGGCCTCCCCTTCATGGTAGCACTAGTTGGCGGGTTGTGGTTGGACTAGGCG

GCTAGGGTATATACCTAGTAGCGGCGGCTGCGGAGTGGAGGGCTGGCGCCCAGCGCGAGGGCGTGGCCTTTCCTCC

TGGACCCGAGAGCGCTCCGCGAGGGACGGCGAGTGAGATAGGCAGCAGCG

10 20 30 40 50 60 MRGSHHHHHH GMASMTGGQQ MGRDLYDDDD KDHPFTACNV ATGPRPPMTT FTGGNKGPAK

       70         80         90        100        110        120 

QQVSLDLRDE GAGMFTSTSP EMRRVVPDDV KGRVKVKVVY VVLEAQYQSA ISAAVKNINA

      130        140        150        160        170        180 

KNSKVCFEVV GYLLEELRDQ KNLDMLKEDV ASANIFIGSL IFIEELAEKI VEAVSPLREK

      190        200        210        220        230        240 

LDACLIFPSM PAVMKLNKLG TFSMAQLGQS KSVFSEFIKS ARKNNDNFEE GLLKLVRTLP

      250        260        270        280        290        300 

KVLKYLPSDK AQDAKNFVNS LQYWLGGNSD NLENLLLNTV SNYVPALKGV DFSVAEPTAY

      310        320        330        340        350        360 

PDVGIWHPLA SGMYEDLKEY LNWYDTRKDM VFAKDAPVIG LVLQRSHLVT GDEGHYSGVV

      370        380        390        400        410        420 

AELESRGAKV IPVFAGGLDF SAPVKKFFYD PLGSGRTFVD TVVSLTGFAL VGGPARQDAP

      430        440        450        460        470        480 

KAIEALKNLN VPYLVSLPLV FQTTEEWLDS ELGVHPVQVA LQVALPELDG AMEPIVFAGR

      490        500        510        520        530        540 

DSNTGKSHSL PDRIASLCAR AVNWANLRKK RNAEKKLAVT VFSFPPDKGN VGTAAYLNVF

      550        560        570        580        590        600 

GSIYRVLKNL QREGYDVGAL PPSEEDLIQS VLTQKEAKFN STDLHIAYKM KVDEYQKLCP

      610        620        630        640        650        660 

YAEALEENWG KPPGTLNTNG QELLVYGRQY GNVFIGVQPT FGYEGDPMRL LFSKSASPHH

      670        680        690        700        710        720 

GFAAYYTFLE KIFKADAVLH FGTHGSLEFM PGKQVGMSGV CYPDSLIGTI PNLYYYAANN

      730        740        750        760        770        780 

PSEATIAKRR SYANTISYLT PPAENAGLYK GLKELKELIS SYQGMRESGR AEQICATIIE

      790        800        810        820        830        840 

TAKLCNLDRD VTLPDADAKD LTMDMRDSVV GQVYRKLMEI ESRLLPCGLH VVGCPPTAEE

      850        860        870        880        890        900 

AVATLVNIAE LDRPDNNPPI KGMPGILARA IGRDIESIYS GNNKGVLADV DQLQRITEAS

      910        920        930        940        950        960 

RTCVREFVKD RTGLNGRIGT NWITNLLKFT GFYVDPWVRG LQNGEFASAN REELITLFNY

      970        980        990       1000       1010       1020 

LEFCLTQVVK DNELGALVEA LNGQYVEPGP GGDPIRNPNV LPTGKNIHAL DPQSIPTQAA

     1030       1040       1050       1060       1070       1080 

LKSARLVVDR LLDRERDNNG GKYPETIALV LWGTDNIKTY GESLAQVMMM VGVKPVADAL

     1090       1100       1110       1120       1130       1140 

GRVNKLEVIP LEELGRPRVD VVVNCSGVFR DLFVNQMLLL DRAIKLAAEQ DEPDEMNFVR

     1150       1160       1170       1180       1190       1200 

KHAKQQAAEL GLQSLRDAAT RVFSNSSGSY SSNVNLAVEN SSWSDESQLQ EMYLKRKSYA

     1210       1220       1230       1240       1250       1260 

FNSDRPGAGG EMQRDVFETA MKTVDVTFQN LDSSEISLTD VSHYFDSDPT KLVASLRNDG

     1270       1280       1290       1300       1310       1320 

RTPNAYIADT TTANAQVRTL GETVRLDART KLLNPKWYEG MLASGYEGVR EIQKRMTNTM

     1330       1340       1350       1360       1370       1380 

GWSATSGMVD NWVYDEANST FIEDAAMAER LMNTNPNSFR KLVATFLEAN GRGYWDAKPE

     1390       1400 

QLERLRQLYM DVEDKIEGVE

References and documentation are available.

Please note the modified algorithm for extinction coefficient.

Number of amino acids: 1400

Molecular weight: 154817.9

Theoretical pI: 5.48

Amino acid composition: Ala (A) 116 8.3% Arg (R) 71 5.1% Asn (N) 79 5.6% Asp (D) 86 6.1% Cys (C) 13 0.9% Gln (Q) 48 3.4% Glu (E) 92 6.6% Gly (G) 105 7.5% His (H) 21 1.5% Ile (I) 51 3.6% Leu (L) 142 10.1% Lys (K) 80 5.7% Met (M) 38 2.7% Phe (F) 51 3.6% Pro (P) 71 5.1% Ser (S) 86 6.1% Thr (T) 71 5.1% Trp (W) 14 1.0% Tyr (Y) 50 3.6% Val (V) 115 8.2% Pyl (O) 0 0.0% Sec (U) 0 0.0% (B) 0 0.0% (Z) 0 0.0% (X) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 178 Total number of positively charged residues (Arg + Lys): 151

Atomic composition:

Carbon C 6872 Hydrogen H 10826 Nitrogen N 1876 Oxygen O 2091 Sulfur S 51

Formula: C6872H10826N1876O2091S51 Total number of atoms: 21716

Extinction coefficients:

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 152250 Abs 0.1% (=1 g/l) 0.983, assuming all pairs of Cys residues form cystines

Ext. coefficient 151500 Abs 0.1% (=1 g/l) 0.979, assuming all Cys residues are reduced

Estimated half-life:

The N-terminal of the sequence considered is M (Met).

The estimated half-life is:

                            30 hours (mammalian reticulocytes, in vitro).
                           >20 hours (yeast, in vivo).
                           >10 hours (Escherichia coli, in vivo).

Instability index:

The instability index (II) is computed to be 32.82 This classifies the protein as stable.

Aliphatic index: 85.87

Grand average of hydropathicity (GRAVY): -0.292

Source

Chlamydomonas reinhardtii

References