Difference between revisions of "R265W"

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(Structure-based Mutation Analysis)
(Side chain properties)
 
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The structures of arginine and tryptophane are very different what can be seen by comparing figures 2 and 3. Moreover the positive charge of arginine gets lost when the hydrophobic tryptophan is introduced. These changes in structure and biochemical properties are quite severe and this mutation is very likely to destroy the protein's function.
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The structures of arginine and tryptophane are very different which can be seen by comparing figures 2 and 3. Moreover the positive charge of arginine gets lost when the hydrophobic tryptophan is introduced. These changes in structure and biochemical properties are quite severe and this mutation is very likely to destroy the protein's function.
   
 
=== Hydrogen Bonding network ===
 
=== Hydrogen Bonding network ===

Latest revision as of 12:10, 26 August 2011

Structure-based Mutation Analysis

Mapping onto crystal structure

Figure 1 shows the protein structure with highlighted amino acids. The mutation position is colored violet, the thiamine pyrophosphate binding sites are orange, and metal binding sites are yellow.

Figure 1: Structure of BCKDHA with highlighted amino acids

This mutation is again quite near by the active site of the protein. Futhermore the most bulky amino acid is introduced in this position which might very likely interfere with the active site and therefore destroy the protein's function.

Side chain properties

Figures 2 and 3 show the superposition of the mutated amino acid with the wildtype. The pictures showing the wildtype structure display the unmutated residue in bold and vice versa.


Figure 2: Structure of arginine on position 265 in the wildtype protein
Figure 3: Structure of tryptophan on position 265 in the mutated protein

The structures of arginine and tryptophane are very different which can be seen by comparing figures 2 and 3. Moreover the positive charge of arginine gets lost when the hydrophobic tryptophan is introduced. These changes in structure and biochemical properties are quite severe and this mutation is very likely to destroy the protein's function.

Hydrogen Bonding network

Hydrogen bonds are interactions between an hydrogen atom and an electronegative atom. Electronegative atoms which often take part in hydrogen bonds are oxygen, nitrogen and fluorine (not present in amino acid side chains). They serve as a hydrogen bond acceptor, whereas a hydrogen bond donor is a electronegative atom bonded to a hydrogen atom. Hydrogen bonds are essential for the three-dimensional structures of proteins. They play a important role in the formation of helices and beta-sheets and cause proteins to fold into a specific structure.

Showing hydrogen bonds with Pymol: A -> find -> polar contacts -> within selection The respective amino acids were colored by element, s.t. oxygen is red, nitrogen is blue, hydrogen is white and sulfur is yellow.

The following figures show the hydrogen bonds between the wildtype residue and its environment compared to the formation of hydrogen bonds when the corresponding residue is mutated.

Figure 4: Hydrogen Bonds for arginine on pos 265 in the wild type structure
Figure 5: Hydrogen Bonds for tryptophan on pos 265 in the mutated type structure

The mutation from arginine to tryptophan leads to a drastic change in the hydrogen bonding network. Arginine, which contains three nitrogen atoms in its side chain is removed and therefore three hydrogen bond acceptors (see Figure 4) are missing in the mutated protein (Figure 5).

foldX Energy Comparison

We used the foldX tool to compare the energy of the wildtype protein and the mutated structure. The following table shows the calculated energy values as well as the percentage of difference, to compare the energy calculations with other tools:

Energy wildtype energy total energy of mutated protein difference
absolute 401.00 460.43 59.43
relative 100% 114% 14%

The total energy of the mutated structure is a higher than the energy of the wildtype protein structure. As protein energies should be low for a stable protein, the increasing energy leads to the assumption that this mutation might be damaging for the protein structure.

minimise Energy Comparison

Next we used the minimise tool to compare the energy of the wildtype protein and the mutated structure. The following table shows the calculated energy values as well as the percentage of difference, to compare the energy calculations with other tools:

Energy wildtype energy total energy of mutated protein difference
absolute -2485.452755 -3989.790625 -1504.33787
relative 100% 62% 38%

The energy calculated by minimise for the mtuated structure is again a lot smaller than the wildtype structure, indicating that the protein structure changed due to the mutation.

gromacs Energy comparison

The Gromacs energy comparison was conducted using the AMBER03 force field. The following table shows the calculated energies for the wildtype protein structure.

Energy Average Err.Est RMSD Tot-Drift (kJ/mol)
Bond 3072.83 2200 -nan -13100.2
Angle 3616.97 230 -nan -1295.57
Potential 2.67001e+07 2.6e+07 -nan -1.60382e+08

Here are the results for the mutated protein structure.

Energy Average Err.Est RMSD Tot-Drift (kJ/mol)
Bond 2473.43 1700 6385.14 -9741.04
Angle 3726.4 330 827.187 1803.54
Potential 5.36e+06 5.3e+06 7.68e+07 -3.26e+07


As we want to compare the Gromacs energies with the other tools, we calculate the ratio of difference considering the potential energy:

Energy wildtype energy total energy of mutated protein difference
absolute 2.67001e+07 5.36e+06 -21340100
relative 100% 20% 80%

The Gromacs energy for the mutated structure is only one fifth of the wildtype-energy. This drastic change in energy shows that the protein's structure changed completely after substituting arginine with tryptophan.

Conclusion

The overall trend of the energy calculation tools stays the same for this mutation. foldX calculates an energy for the mutated structure a little bit higher than the wildtype, while minimise and gromacs return smaller energies for the mutated proteins. This is different compared to the C264W mutation, although these mutations are right next to each other and each time the wildtype amino acid is substituted by tryptophan. Comparing these two mutations and the corresponding energy calculation results we would conclude that the effect of the C264W mutation is totally different than the effect of this mutation.

return to Structure-based mutation analysis