Team:SVCE CHENNAI/Description
In-silico Results
In-silico studies play a major part of any biotechnological project.
Part Enhancement 2 : cjBlue,green chromoprotein
Similar insilico analysis to tsPurple was done for this part. The sequence for this part was obtained from the iGEM registry and submitted to SWISS MODEL . The protein with the highest sequence similarity was found to be template 2ib5.1.A, which is also a chromoprotein. The secondary structure of this protein was used to construct a 3D model for cjBlue. The predicted 3D structure was a homooctamer. Each monomer subunit had a ? barrel with a central ? helix. To determine if the predicted model was authentic, we generated the Ramachandran Plot for this model by using the SWISS MODEL Workspace. A score of 94% in the most favoured regions as was seen in the Ramachandran plot showed that the predicted model of cjBlue chromoprotein is a very good prediction.
Physical and chemical properties
Number of amino acids: 232
Molecular weight: 26173.80
Theoretical pI: 7.53
Total number of negatively charged residues (Asp + Glu): 24
Total number of positively charged residues (Arg + Lys): 25
Atomic composition:
Carbon (C) - 1151
Hydrogen (H) - 1776
Nitrogen (N) - 314
Oxygen (O) - 347
Sulfur (S) - 19
Formula: C1151H1776N314O347S19
Total number of atoms: 3607
Extinction coefficients:
Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.
Ext. coefficient : 28015
Abs 0.1% (=1 g/l): 1.070, assuming all pairs of Cys residues form cystines
Ext. coefficient : 27390
Abs 0.1% (=1 g/l): 1.046, 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 29.86.
This classifies the protein as stable.
Aliphatic index: 58.84
Grand average of hydropathicity (GRAVY): -0.558
Interested in knowing about this protein? click here.
Part Enhancement 3 : RNA thermometer (ROSE)
We used SimRNAweb which is a web server for RNA 3D structure modeling to create a model for the RNA thermometer.
Interested in seeing the secondary structure of this model? Click here.
Part Enhancement 1 : tsPurple chromoprotein
Insilico analysis was done to construct a 3D model for this part. The sequence for this part was obtained from the iGEM registry and submitted to SWISS MODEL . The protein with the highest sequence similarity was found to be FAR-RED FLUORESCENT PROTEIN AQ143. The secondary structure of this protein was used to construct a 3D model for tsPurple. The predicted 3D structure was a homotetramer. Each monomer subunit had a β barrel with a central α helix.
To determine if the predicted model was authentic, we generated the Ramachandran Plot for this model by using the SWISS MODEL Workspace. A score of 93.3% in the most favoured regions as was seen in the Ramachandran plot showed that the predicted model of Tinsel Purple chromoprotein is a very good prediction.
To find further information on this protein we used ExPASy - ProtParam tool to compute the various physical and chemical parameters for tsPurple. The computed parameters included the molecular weight, theoretical pI, amino acid composition, atomic composition, extinction coefficient, estimated half-life, instability index, aliphatic index and grand average of hydropathicity (GRAVY).
And finally, it would be a shame if you created a wonderful construct with this part, but then this protein was cleaved by an enzyme produced by your chassis organism, now wouldn't it? To tackle this problem we created a database of all the enzymes that would cleave this chromoprotein, the number and position of the cleavage sites using ExPASy Peptide Cutter.
Physical and chemical properties
Number of amino acids: 228
Molecular weight: 25546.26
Theoretical pI: 6.65
Total number of negatively charged residues (Asp + Glu): 27
Total number of positively charged residues (Arg + Lys): 26
Atomic composition:
Carbon (C) - 1134
Hydrogen (H) - 1747
Nitrogen (N) - 303
Oxygen (O) - 334
Sulfur (S) - 18
Formula: C1134H1747N303O334S18
Total number of atoms: 3536
Extinction coefficients:
Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.
Ext. coefficient : 24910
Abs 0.1% (=1 g/l): 0.975, assuming all pairs of Cys residues form cystines
Ext. coefficient : 24410
Abs 0.1% (=1 g/l): 0.956, 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 31.19
This classifies the protein as stable.
Aliphatic index: 61.97
Grand average of hydropathicity (GRAVY): -0.395
Interested in knowing about this protein? click here.
