Difference between revisions of "Part:BBa K2740016"

Latest revision as of 10:15, 8 October 2018

CR1 nifE

CR1 nifE encodes the cofactor NifE for the maturation of functional molybdenum-iron protein.

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
INCOMPATIBLE WITH RFC[25]
Illegal NgoMIV site found at 6
Illegal AgeI site found at 379
1000
COMPATIBLE WITH RFC[1000]

Parameter of Protein

Number of amino acids: 453

Molecular weight: 49636.54

Theoretical pI: 6.00

Amino acid composition:
Ala (A) 39 8.6%
Arg (R) 29 6.4%
Asn (N) 11 2.4%
Asp (D) 19 4.2%
Cys (C)   7 1.5%
Gln (Q) 16 3.5%
Glu (E) 37 8.2%
Gly (G) 45 9.9%
His (H) 10 2.2%
Ile (I)    30 6.6%
Leu (L) 38 8.4%
Lys (K) 21 4.6%
Met (M) 11 2.4%
Phe (F) 12 2.6%
Pro (P) 21   4.6%
Ser (S) 32   7.1%
Thr (T) 24 5.3%
Trp (W) 6   1.3%
Tyr (Y) 15 3.3%
Val (V) 30 6.6%
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): 56
Total number of positively charged residues (Arg + Lys): 50

Atomic composition:

Carbon      C          2192
Hydrogen    H         3480
Nitrogen    N            614
Oxygen      O          664
Sulfur      S              18

Formula: C2192H3480N614O664S18
Total number of atoms: 6968

Extinction coefficients:

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

Ext. coefficient 55725
Abs 0.1% (=1 g/l) 1.123, assuming all pairs of Cys residues form cystines

Ext. coefficient 55350
Abs 0.1% (=1 g/l) 1.115, 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 40.63
This classifies the protein as unstable.

Aliphatic index: 86.36

Grand average of hydropathicity (GRAVY): -0.234

Design Notes

Nitrogenase is a complex enzyme system consisting of nine protein components. Additionally, to maintain stoichiometry of these protein components is an essential requirement for nitrogenase biosynthesis and activity. However, there is only one copy of each structure gene present in the nif gene cluster. Therefore, cloning each of these nif genes and setting as independent part can facilitate the regulation of balancing expression ratios from the transcription and/or translation level(s) when they are heterogeneously expressed in non-diazotrophic hosts.

@@ Line 17: / Line 17: @@
 <partinfo>BBa_K2740016 parameters</partinfo>
 <!-- -->
-Parameter of Protein
+<h2>Parameter of Protein </h2>
+<p align="left">Number  of amino acids: 453</p>
-Number of amino acids: 453
+<p align="left">Molecular  weight: 49636.54</p>
+<p align="left">Theoretical  pI: 6.00</p>
-Molecular weight: 49636.54
+<p align="left">Amino  acid composition: <br />
+  Ala  (A)  39    8.6%<br />
-Theoretical pI: 6.00
+  Arg  (R)  29    6.4%<br />
+  Asn  (N)  11   2.4%<br />
-Amino acid composition:
+  Asp  (D)  19   4.2%<br />
-Ala (A)  39	  8.6%
+  Cys  (C)   7    1.5%<br />
-Arg (R)  29	  6.4%
+  Gln  (Q)  16    3.5%<br />
-Asn (N)  11	  2.4%
+  Glu  (E)  37    8.2%<br />
-Asp (D)  19	  4.2%
+  Gly  (G)  45    9.9%<br />
-Cys (C)   7	  1.5%
+  His  (H)  10    2.2%<br />
-Gln (Q)  16	  3.5%
+  Ile  (I)    30   6.6%<br />
-Glu (E)  37	  8.2%
+  Leu  (L)  38    8.4%<br />
-Gly (G)  45	  9.9%
+  Lys  (K)  21    4.6%<br />
-His (H)  10	  2.2%
+  Met  (M)  11   2.4%<br />
-Ile (I)    30	  6.6%
+  Phe  (F)  12    2.6%<br />
-Leu (L)  38	  8.4%
+  Pro  (P)  21     4.6%<br />
-Lys (K)  21	  4.6%
+  Ser (S)  32     7.1%<br />
-Met (M)  11	  2.4%
+  Thr  (T)  24    5.3%<br />
-Phe (F)  12	  2.6%
+  Trp  (W)  6     1.3%<br />
-Pro (P)  21	  4.6%
+  Tyr  (Y)  15    3.3%<br />
-Ser (S)  32	  7.1%
+  Val  (V)  30    6.6%<br />
-Thr (T)  24	  5.3%
+  Pyl  (O)   0     0.0%<br />
-Trp (W)  6	  1.3%
+  Sec  (U)   0    0.0%</p>
-Tyr (Y)  15	  3.3%
+<p align="left"> (B)   0          0.0%<br />
-Val (V)  30	  6.6%
+  (Z)   0   0.0%<br />
-Pyl (O)   0	  0.0%
+  (X)   0          0.0%</p>
-Sec (U)   0	  0.0%
+<p align="left">&nbsp;</p>
+<p align="left">Total  number of negatively charged residues (Asp + Glu): 56<br />
- (B)   0	  0.0%
+  Total  number of positively charged residues (Arg + Lys): 50</p>
- (Z)   0	  0.0%
+<p align="left">Atomic  composition:</p>
- (X)   0	  0.0%
+<p align="left">Carbon      C          2192<br />
+  Hydrogen    H         3480<br />
+  Nitrogen    N            614<br />
-Total number of negatively charged residues (Asp + Glu): 56
+  Oxygen      O          664<br />
-Total number of positively charged residues (Arg + Lys): 50
+  Sulfur      S              18</p>
+<p align="left">Formula:  C2192H3480N614O664S18<br />
-Atomic composition:
+  Total  number of atoms: 6968</p>
+<p align="left">Extinction  coefficients:</p>
-Carbon      C	      2192
+<p align="left">Extinction  coefficients are in units of  M-1 cm-1,  at 280 nm measured in water.</p>
-Hydrogen    H	      3480
+<p align="left">Ext.  coefficient    55725<br />
-Nitrogen    N	       614
+  Abs  0.1% (=1 g/l)   1.123, assuming all pairs  of Cys residues form cystines</p>
-Oxygen      O	       664
+<p align="left">&nbsp;</p>
-Sulfur      S	        18
+<p align="left">Ext.  coefficient    55350<br />
+  Abs  0.1% (=1 g/l)   1.115, assuming all Cys  residues are reduced</p>
-Formula: C2192H3480N614O664S18
+<p align="left">Estimated  half-life:</p>
-Total number of atoms: 6968
+<p align="left">The  N-terminal of the sequence considered is M (Met).</p>
+<p align="left">The  estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).<br />
-Extinction coefficients:
+  &gt;20 hours  (yeast, in vivo).<br />
+  &gt;10 hours (Escherichia  coli, in vivo).</p>
-Extinction coefficients are in units of  M-1 cm-1, at 280 nm measured in water.
+<p align="left">&nbsp;</p>
+<p align="left">Instability  index:</p>
-Ext. coefficient    55725
+<p align="left">The  instability index (II) is computed to be 40.63<br />
-Abs 0.1% (=1 g/l)   1.123, assuming all pairs of Cys residues form cystines
+  This  classifies the protein as unstable.</p>
+<p align="left">&nbsp;</p>
+<p align="left">Aliphatic  index: 86.36</p>
-Ext. coefficient    55350
+<p align="left">Grand  average of hydropathicity (GRAVY): -0.234</p>
-Abs 0.1% (=1 g/l)   1.115, assuming all Cys residues are reduced
+<div>
+  <h2>Design Notes</h2>
-Estimated half-life:
+</div>
+<p align="left">Nitrogenase  is a complex enzyme system consisting of nine protein components. Additionally,  to maintain stoichiometry of these protein components is an essential  requirement for nitrogenase biosynthesis and activity. However, there is only  one copy of each structure gene present  in the nif gene cluster. Therefore, cloning each of these nif genes and setting  as independent part can facilitate the regulation of balancing expression  ratios from the transcription and/or translation level(s) when they are heterogeneously expressed in non-diazotrophic hosts.</p>
-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 40.63
-This classifies the protein as unstable.
-Aliphatic index: 86.36
-Grand average of hydropathicity (GRAVY): -0.234
-Design Notes
-Nitrogenase is a complex enzyme system consisting of nine protein components. Additionally, to maintain stoichiometry of these protein components is an essential requirement for nitrogenase biosynthesis and activity. However, there is only one copy of each structure gene present in the nif gene cluster. Therefore, cloning each of these nif genes and setting as independent part can facilitate the regulation of balancing expression ratios from the transcription and/or translation level(s) when they are heterogeneously expressed in non-diazotrophic hosts.We sent the sequences of the PCR template to synthesis, but unfortunately, EcoRI and PstI striction enzyme cut site was involved after they promoted it again. But the part can be manipulated by XbaI and SpeI or can be assembled by gibson assembly,that is what we did.