Difference between revisions of "Part:BBa K2993003"
 
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This part is special in its own way because it actually is a fusion of one half of a protein to a whole array of zinc-finger proteins. Much like a multi fusion protein.  
 
This part is special in its own way because it actually is a fusion of one half of a protein to a whole array of zinc-finger proteins. Much like a multi fusion protein.  
  
−
The full sequence of this part is:
+
The full sequence of this part is (we tried fitting it in the page width):
 +
 
 
GAATTCGCGGCCGCTTCTAGAGGTTGACGGCTAGCTCAGTCCTAGGTACAG
 
GAATTCGCGGCCGCTTCTAGAGGTTGACGGCTAGCTCAGTCCTAGGTACAG
 
TGCTAGCTtcaagaGAAAGAGGGGACAAaCTAGATGGCCGGCCTGGAACCAGGCGAAAAA
 
TGCTAGCTtcaagaGAAAGAGGGGACAAaCTAGATGGCCGGCCTGGAACCAGGCGAAAAA
Line 29: Line 30:
 
cgattaaatgctgactcagtattgtgggggggccataaagttttcatggtgaaacctgaagagccttttcagccagtt
 
cgattaaatgctgactcagtattgtgggggggccataaagttttcatggtgaaacctgaagagccttttcagccagtt
 
aaggaagcgactcaactcatgaatACCGGCCACCACCATCACCATCACCATCACTACTAGTAGCGGCCGCTGCAG
 
aaggaagcgactcaactcatgaatACCGGCCACCACCATCACCATCACCATCACTACTAGTAGCGGCCGCTGCAG
 +
 +
 +
 +
Even though the protein has not been brought to expression, theoretical estimations were made with known software from ExPASy (https://web.expasy.org/protparam/) as shown below:
 +
 +
ProtParam
 +
User-provided sequence:
 +
        10        20        30        40        50        60
 +
MAGLEPGEKP YKCPECGKSF SQSGHLTEHQ RTHTGEKPYK CPECGKSFSR EDNLHTHQRT
 +
 +
        70        80        90        100        110        120
 +
HTGEKPYKCP ECGKSFSSKK HLAEHQRTHT GEKPYKCPEC GKSFSRTDTL RDHQRTHTGE
 +
 +
      130        140        150        160        170        180
 +
KPYKCPECGK SFSREDNLHT HQRTHTGEKP YKCPECGKSF SREDNLHTHQ RTHTGKKTST
 +
 +
      190        200        210        220        230        240
 +
GGGSAGGSGS GSSGGSSGAS GTGTAGGTGS GSGTGSGTGK SMSSMVSDTS CTFPSSDGIF
 +
 +
      250        260        270        280        290        300
 +
WKHWIQTKDG QCGSPLVSTR DGFIVGIHSA SNFTNTNNYF TSVPKNFMEL LTNQEAQQWV
 +
 +
      310        320        330        340
 +
SGWRLNADSV LWGGHKVFMV KPEEPFQPVK EATQLMNTGH HHHHHHHY
 +
 +
 +
References and documentation are available.
 +
 +
Number of amino acids: 348
 +
 +
Molecular weight: 38135.93
 +
 +
Theoretical pI: 8.83
 +
 +
Amino acid composition:
 +
Ala (A)  9   2.6%
 +
Arg (R)  13   3.7%
 +
Asn (N)  11   3.2%
 +
Asp (D)  10   2.9%
 +
Cys (C)  14   4.0%
 +
Gln (Q)  14   4.0%
 +
Glu (E)  23   6.6%
 +
Gly (G)  44 12.6%
 +
His (H)  28   8.0%
 +
Ile (I)  4   1.1%
 +
Leu (L)  13   3.7%
 +
Lys (K)  29   8.3%
 +
Met (M)  6   1.7%
 +
Phe (F)  14   4.0%
 +
Pro (P)  19   5.5%
 +
Ser (S)  40 11.5%
 +
Thr (T)  35 10.1%
 +
Trp (W)  5   1.4%
 +
Tyr (Y)  8   2.3%
 +
Val (V)  9   2.6%
 +
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): 33
 +
Total number of positively charged residues (Arg + Lys): 42
 +
 +
Atomic composition:
 +
 +
Carbon      C       1631
 +
Hydrogen    H       2486
 +
Nitrogen    N       502
 +
Oxygen      O       523
 +
Sulfur      S         20
 +
 +
Formula: C1631H2486N502O523S20
 +
Total number of atoms: 5162
 +
 +
Extinction coefficients:
 +
 +
Extinction coefficients are in units of  M-1 cm-1, at 280 nm measured in water.
 +
 +
Ext. coefficient    40295
 +
Abs 0.1% (=1 g/l)  1.057, assuming all pairs of Cys residues form cystines
 +
 +
 +
Ext. coefficient    39420
 +
Abs 0.1% (=1 g/l)  1.034, 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 33.56
 +
This classifies the protein as stable.
 +
 +
 +
 +
Aliphatic index: 29.14
 +
 +
Grand average of hydropathicity (GRAVY): -1.082
 +
  
 
<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here

Latest revision as of 19:03, 20 October 2019


Fusion protein of ARRAY2-GSAT-C-TEV-6xHisTag

This part contains the C-TEV split protein fused to zincfinger array 2 via a GSAT-linker.

This fusion protein should be used with its counterpart which contains the C-TEV protein fused to another zinc-finger array with a binding affinity for a different DNA sequence. On our iGEMteam_Rotterdam2019 webpage, you can read all about how to use this part properly.

This part is special in its own way because it actually is a fusion of one half of a protein to a whole array of zinc-finger proteins. Much like a multi fusion protein.

The full sequence of this part is (we tried fitting it in the page width):

GAATTCGCGGCCGCTTCTAGAGGTTGACGGCTAGCTCAGTCCTAGGTACAG TGCTAGCTtcaagaGAAAGAGGGGACAAaCTAGATGGCCGGCCTGGAACCAGGCGAAAAA CCGTACAAATGCCCGGAATGTGGTAAAAGCTTCAGCCAGAGCGGCCATCTG ACCGAACACCAGCGTACGCATACCGGCGAAAAACCATATAAATGTCCGGAATGTGGCAAA TCTTTTAGCCGTGAAGATAACCTTCACACCCATCAGCGCACCCATACTGGTG AAAAACCGTATAAGTGCCCGGAATGTGGCAAAAGTTTTAGCAGCAAGAAACATCTGGCG GAACATCAACGCACCCACACCGGCGAAAAGCCGTATAAATGCCCGGAATGCG GCAAAAGCTTTAGCCGCACCGATACCCTGCGCGATCACCAACGCACTCACACAGGTGA GAAACCCTACAAGTGCCCAGAATGTGGAAAATCATTTAGCCGCGAGGATAAT TTACACACCCATCAACGTACGCATACAGGCGAAAAGCCATACAAGTGTCCCGAGTGTGG CAAAAGTTTCTCGCGCGAAGATAATTTGCATACGCACCAACGCACGCATACT GGCAAGAAGACAAGTACCGGCGGAGGGTCGGCAGGAGGGAGTGGTTCCGGATCTAGTG GTGGAAGCTCAGGAGCTAGTGGTACAGGCACTGCGGGTGGCACTGGCAGCG GTTCCGGTACTGGCTCTGGCACCGGCaagagcatgtctagcatggtgtcagacactagctgcacatt cccttcatctgatggcatattctggaagcattggattcaaaccaaggatgggcagtgtggcagtccat tagtatcaactagagatgggttcattgttggtatacactcagcatcgaatttcaccaacacaaacaattatttcacaa gcgtgccgaaaaacttcatggaattgttgacaaatcaggaggcgcagcagtgggttagtggttgg cgattaaatgctgactcagtattgtgggggggccataaagttttcatggtgaaacctgaagagccttttcagccagtt aaggaagcgactcaactcatgaatACCGGCCACCACCATCACCATCACCATCACTACTAGTAGCGGCCGCTGCAG


Even though the protein has not been brought to expression, theoretical estimations were made with known software from ExPASy (https://web.expasy.org/protparam/) as shown below:

ProtParam User-provided sequence: 

       10         20         30         40         50         60

MAGLEPGEKP YKCPECGKSF SQSGHLTEHQ RTHTGEKPYK CPECGKSFSR EDNLHTHQRT 

       70         80         90        100        110        120

HTGEKPYKCP ECGKSFSSKK HLAEHQRTHT GEKPYKCPEC GKSFSRTDTL RDHQRTHTGE 

      130        140        150        160        170        180

KPYKCPECGK SFSREDNLHT HQRTHTGEKP YKCPECGKSF SREDNLHTHQ RTHTGKKTST 

      190        200        210        220        230        240

GGGSAGGSGS GSSGGSSGAS GTGTAGGTGS GSGTGSGTGK SMSSMVSDTS CTFPSSDGIF 

      250        260        270        280        290        300

WKHWIQTKDG QCGSPLVSTR DGFIVGIHSA SNFTNTNNYF TSVPKNFMEL LTNQEAQQWV 

      310        320        330        340

SGWRLNADSV LWGGHKVFMV KPEEPFQPVK EATQLMNTGH HHHHHHHY


References and documentation are available.

Number of amino acids: 348

Molecular weight: 38135.93

Theoretical pI: 8.83

Amino acid composition: Ala (A) 9 2.6% Arg (R) 13 3.7% Asn (N) 11 3.2% Asp (D) 10 2.9% Cys (C) 14 4.0% Gln (Q) 14 4.0% Glu (E) 23 6.6% Gly (G) 44 12.6% His (H) 28 8.0% Ile (I) 4 1.1% Leu (L) 13 3.7% Lys (K) 29 8.3% Met (M) 6 1.7% Phe (F) 14 4.0% Pro (P) 19 5.5% Ser (S) 40 11.5% Thr (T) 35 10.1% Trp (W) 5 1.4% Tyr (Y) 8 2.3% Val (V) 9 2.6% 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): 33 Total number of positively charged residues (Arg + Lys): 42

Atomic composition:

Carbon C 1631 Hydrogen H 2486 Nitrogen N 502 Oxygen O 523 Sulfur S 20

Formula: C1631H2486N502O523S20 Total number of atoms: 5162

Extinction coefficients:

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

Ext. coefficient 40295 Abs 0.1% (=1 g/l) 1.057, assuming all pairs of Cys residues form cystines


Ext. coefficient 39420 Abs 0.1% (=1 g/l) 1.034, 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 33.56 This classifies the protein as stable.


Aliphatic index: 29.14

Grand average of hydropathicity (GRAVY): -1.082


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 672
    Illegal SapI.rc site found at 982