Difference between revisions of "Part:BBa K1080002:Design"

(→‎Design Notes)
Line 16: Line 16:
 
<b>Amino acid sequence:</b>
 
<b>Amino acid sequence:</b>
  
−
<FONT FACE="courier"> MGRDLYDDDD KDHPFTLRAM KVSEEDSKGF DADVSTRLAR
+
<FONT FACE="courier"> MRAMKVSEED SKGFDADVST RLARSYPLAA VVGQDNIKQA LLLGAVDTGL GGIAIAGRRG<br> KSIMARGL HALLPPIEVV EGSICNADPE DPRSWEAGLA EKYAGGPVKT KMRSAPFVQI<br> DGVNVVEREG ISISHPCRPL LIATYNPEEG PLREHLLDRI AIGLSADVPS TSDERVKAID<br> AAIRFQDKPQ DTIDDTAELT DALRTSVILA REYLKDVTIA PEQVTYIVEE ARRGGVQGHR<br> AELYAVKCAK ACAALEGRER VNKDDLRQAV QLVILPRATI LDQPPPEQEQ PPPPPPPPPP<br> PPPQDQMEDE DQEEKEDEKE EEEKENEDQD EPEIPQEFMF ESEGVIMDPS ILMFAQQQQR<br> AQGRSGRAKT LIFSDDRGRY IKPMLPKGDK VKRLAVDATL RAAAPYQKIR RQQAISEGKV<br> QRKVYVDKPD MRSKKLARKA GALVIFVVDA SGSMALNRMS AAKGACMRLL AESYTSRDQV<br>  VMMVLITDGR ANVSLAKSNE DPEALKPDAP KPTADSLKDE VRDMAKKAAS AGINVLVIDT<br> ENKFVSTGFA EEISKAAQGK YYYLPNASDA AIAAAASGAM AAAKGGY </FONT>
−
 
+
 
−
SYPLAAVVGQ DNIKQALLLG AVDTGLGGIA IAGRRGTAKS IMARGLHALL PPIEVVEGSI
+
References and documentation are available.  
−
 
+
−
CNADPEDPRS WEAGLAEKYA GGPVKTKMRS APFVQIPLGV TEDRLVGTVD IEASMKEGKT
+
−
 
+
−
VFQPGLLAEA HRGILYVDEI NLLDDGIANL LLSILSDGVN VVEREGISIS HPCRPLLIAT
+
−
 
+
−
YNPEEGPLRE HLLDRIAIGL SADVPSTSDE RVKAIDAAIR FQDKPQDTID DTAELTDALR
+
−
 
+
−
TSVILAREYL KDVTIAPEQV TYIVEEARRG GVQGHRAELY AVKCAKACAA LEGRERVNKD
+
−
 
+
−
DLRQAVQLVI LPRATILDQP PPEQEQPPPP PPPPPPPPPQ DQMEDEDQEE KEDEKEEEEK
+
−
 
+
−
ENEDQDEPEI PQEFMFESEG VIMDPSILMF AQQQQRAQGR SGRAKTLIFS DDRGRYIKPM
+
−
 
+
−
LPKGDKVKRL AVDATLRAAA PYQKIRRQQA ISEGKVQRKV YVDKPDMRSK KLARKAGALV
+
−
 
+
−
IFVVDASGSM ALNRMSAAKG ACMRLLAESY TSRDQVCLIP FYGDKAEVLL PPSKSIAMAR
+
−
+
−
RRLDSLPCGG GSPLAHGLST AVRVGMQASQ AGEVGRVMMV LITDGRANVS LAKSNEDPEA
+
−
 
+
−
LKPDAPKPTA DSLKDEVRDM AKKAASAGIN VLVIDTENKF VSTGFAEEIS KAAQGKYYYL
+
−
 
+
−
PNASDAAIAA AASGAMAAAK GGY </FONT>
+
−
 
+
−
 
+
−
References and documentation are available.
+
−
 
+
 
Please note the modified algorithm for extinction coefficient.
 
Please note the modified algorithm for extinction coefficient.
  
 +
--------------------------------------------------------------------------------
 +
Number of amino acids: 707
  
−
Number of amino acids: 743
+
Molecular weight: 76420.1
  
−
Molecular weight: 80527.5
+
Theoretical pI: 5.23
  
−
Theoretical pI: 5.37
+
Amino acid composition: Ala (A)  88 12.4%
 +
Arg (R)  47   6.6%
 +
Asn (N)  14   2.0%
 +
Asp (D)  51   7.2%
 +
Cys (C)  7   1.0%
 +
Gln (Q)  31   4.4%
 +
Glu (E)  55   7.8%
 +
Gly (G)  51   7.2%
 +
His (H)  6   0.8%
 +
Ile (I)  40   5.7%
 +
Leu (L)  60   8.5%
 +
Lys (K)  43   6.1%
 +
Met (M)  20   2.8%
 +
Phe (F)  12   1.7%
 +
Pro (P)  50   7.1%
 +
Ser (S)  42   5.9%
 +
Thr (T)  24   3.4%
 +
Trp (W)  1   0.1%
 +
Tyr (Y)  16   2.3%
 +
Val (V)  49   6.9%
 +
Pyl (O)  0   0.0%
 +
Sec (U)  0   0.0%
  
−
Amino acid composition:
+
(B)   0   0.0%
−
Ala (A) 89 12.0% Arg (R) 49 6.6% Asn (N) 14 1.9% Asp (D) 57 7.7% Cys (C) 7 0.9% Gln (Q) 33 4.4% Glu (E) 55 7.4% Gly (G) 56 7.5% His (H) 13 1.7% Ile (I) 40 5.4% Leu (L) 62 8.3% Lys (K) 44 5.9% Met (M) 23 3.1% Phe (F) 13 1.7% Pro (P) 51 6.9% Ser (S) 44 5.9% Thr (T) 26 3.5% Trp (W) 1 0.1% Tyr (Y) 17 2.3% Val (V) 49 6.6% Pyl (O) 0 0.0% Sec (U) 0 0.0% (B) 0 0.0% (Z) 0 0.0% (X) 0 0.0%
+
(Z)   0   0.0%
 +
(X)   0   0.0%
  
  
−
Total number of negatively charged residues (Asp + Glu): 112
+
Total number of negatively charged residues (Asp + Glu): 106
−
Total number of positively charged residues (Arg + Lys): 93
+
Total number of positively charged residues (Arg + Lys): 90
  
 
Atomic composition:
 
Atomic composition:
  
−
Carbon      C       3504
+
Carbon      C       3333
−
Hydrogen    H       5684
+
Hydrogen    H       5435
−
Nitrogen    N       1008
+
Nitrogen    N       949
−
Oxygen      O       1102
+
Oxygen      O       1047
−
Sulfur      S         30
+
Sulfur      S         27
  
−
Formula: C3504H5684N1008O1102S30
+
Formula: C3333H5435N949O1047S27
−
Total number of atoms: 11328
+
Total number of atoms: 10791
  
 
Extinction coefficients:
 
Extinction coefficients:
Line 76: Line 74:
 
Extinction coefficients are in units of  M-1 cm-1, at 280 nm measured in water.
 
Extinction coefficients are in units of  M-1 cm-1, at 280 nm measured in water.
  
−
Ext. coefficient    31205
+
Ext. coefficient    29715
−
Abs 0.1% (=1 g/l)  0.388, assuming all pairs of Cys residues form cystines
+
Abs 0.1% (=1 g/l)  0.389, assuming all pairs of Cys residues form cystines
  
  
−
Ext. coefficient    30830
+
Ext. coefficient    29340
−
Abs 0.1% (=1 g/l)  0.383, assuming all Cys residues are reduced
+
Abs 0.1% (=1 g/l)  0.384, assuming all Cys residues are reduced
  
 
Estimated half-life:
 
Estimated half-life:
Line 87: Line 85:
 
The N-terminal of the sequence considered is M (Met).
 
The N-terminal of the sequence considered is M (Met).
  
−
The estimated half-life is:
+
The estimated half-life is:  
 
                             30 hours (mammalian reticulocytes, in vitro).
 
                             30 hours (mammalian reticulocytes, in vitro).
 
                             >20 hours (yeast, in vivo).
 
                             >20 hours (yeast, in vivo).
Line 95: Line 93:
 
Instability index:
 
Instability index:
  
−
The instability index (II) is computed to be 44.71
+
The instability index (II) is computed to be 45.56
 
This classifies the protein as unstable.
 
This classifies the protein as unstable.
  
  
  
−
Aliphatic index: 84.64
+
Aliphatic index: 87.71
−
 
+
−
Grand average of hydropathicity (GRAVY): -0.392
+
−
 
+
−
===Source===
+
−
 
+
−
Chlamydomonas reinhardtii
+
  
−
===References===
+
Grand average of hydropathicity (GRAVY): -0.337

Revision as of 02:26, 26 September 2013

ChlD


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NotI site found at 1334
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 2039
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 614
    Illegal NgoMIV site found at 1433
    Illegal NgoMIV site found at 1975
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

Incorporated sequence overlap for Gibson assembly and no GC rich region or restriction site in sequence


ChlD Clone: DNA sequence from translation start site:


Amino acid sequence:

MRAMKVSEED SKGFDADVST RLARSYPLAA VVGQDNIKQA LLLGAVDTGL GGIAIAGRRG
KSIMARGL HALLPPIEVV EGSICNADPE DPRSWEAGLA EKYAGGPVKT KMRSAPFVQI
DGVNVVEREG ISISHPCRPL LIATYNPEEG PLREHLLDRI AIGLSADVPS TSDERVKAID
AAIRFQDKPQ DTIDDTAELT DALRTSVILA REYLKDVTIA PEQVTYIVEE ARRGGVQGHR
AELYAVKCAK ACAALEGRER VNKDDLRQAV QLVILPRATI LDQPPPEQEQ PPPPPPPPPP
PPPQDQMEDE DQEEKEDEKE EEEKENEDQD EPEIPQEFMF ESEGVIMDPS ILMFAQQQQR
AQGRSGRAKT LIFSDDRGRY IKPMLPKGDK VKRLAVDATL RAAAPYQKIR RQQAISEGKV
QRKVYVDKPD MRSKKLARKA GALVIFVVDA SGSMALNRMS AAKGACMRLL AESYTSRDQV
VMMVLITDGR ANVSLAKSNE DPEALKPDAP KPTADSLKDE VRDMAKKAAS AGINVLVIDT
ENKFVSTGFA EEISKAAQGK YYYLPNASDA AIAAAASGAM AAAKGGY

References and documentation are available. Please note the modified algorithm for extinction coefficient.

Number of amino acids: 707

Molecular weight: 76420.1

Theoretical pI: 5.23

Amino acid composition: Ala (A) 88 12.4% Arg (R) 47 6.6% Asn (N) 14 2.0% Asp (D) 51 7.2% Cys (C) 7 1.0% Gln (Q) 31 4.4% Glu (E) 55 7.8% Gly (G) 51 7.2% His (H) 6 0.8% Ile (I) 40 5.7% Leu (L) 60 8.5% Lys (K) 43 6.1% Met (M) 20 2.8% Phe (F) 12 1.7% Pro (P) 50 7.1% Ser (S) 42 5.9% Thr (T) 24 3.4% Trp (W) 1 0.1% Tyr (Y) 16 2.3% Val (V) 49 6.9% 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): 106 Total number of positively charged residues (Arg + Lys): 90

Atomic composition:

Carbon C 3333 Hydrogen H 5435 Nitrogen N 949 Oxygen O 1047 Sulfur S 27

Formula: C3333H5435N949O1047S27 Total number of atoms: 10791

Extinction coefficients:

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

Ext. coefficient 29715 Abs 0.1% (=1 g/l) 0.389, assuming all pairs of Cys residues form cystines


Ext. coefficient 29340 Abs 0.1% (=1 g/l) 0.384, 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 45.56 This classifies the protein as unstable.


Aliphatic index: 87.71

Grand average of hydropathicity (GRAVY): -0.337