F409C

Structure-based Mutation Analysis

Side chain properties

Figure 2: Structure of phenylalanine on position 409 in the wildtype protein

Figure 3: Structure of cysteine on position 409 in the mutated protein

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 phenylalanine on pos 409 in the wild type structure

Figure 5: Hydrogen Bonds for cysteine on pos 409 in the mutated type structure

The phenylalanine side chain in the wildtype protein does not participate in any hydrogen bonds (see Figure 4). The mutation to serine doesn't introduce new hydrogen bonding donors or acceptors, therefore the mutation has no effect on the hydrogen bonding network (see 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	433.33	32.33
relative	100%	108%	8%

The total energy of the mutated structure is a little bit 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	-4358.528143	-1873.075388
relative	100%	57%	43%

The mutated structure has an energy that is much smaller than the wildtype

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	2341.69	1600	6048.14	-9087.07
Angle	3597.89	240	594.267	-1309.54
Potential	4.68e+06	4.7e+06	7.12e+07	-2.85e+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	4.68e+06	-22020100
relative	100%	17%	83%

The calculated energy for the mutated structure is much smaller than for the wildtype structure.

Conclusion

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