Gaucher Disease: Task 08 - Sequence-based mutation analysis
Contents
Mutation Set
<figtable id="sele">
Mutations | |||||
---|---|---|---|---|---|
mRNA | Protein | ||||
Reference | Sequence Position | Codon change | Codon Number | Amino Acid change | One letter code |
rs368786234 | 656 | AGC ⇒ AGA | 77 | Ser ⇒ Arg | S77R |
rs374003673 | 847 | AAT ⇒ AGT | 141 | Asn ⇒ Ser | N141S |
CM880035 | - | CGG ⇒ CAG | 159 | Arg ⇒ Gln | R159Q |
rs374591570 | 1062 | CTC ⇒ TTC | 213 | Leu ⇒ Phe | L213F |
CM992894 | - | GGA ⇒ GAA | 241 | Gly ⇒ Glu | G241E |
rs371083513 | 1470 | GTA ⇒ ATA | 349 | Val ⇒ Ile | V349I |
CM960697 | - | ACG ⇒ ATG | 408 | Thr ⇒ Met | T408M |
CM880036 | - | AAC ⇒ AGC | 409 | Asn ⇒ Ser | N409S |
CM870010 | - | CTG ⇒ CCG | 483 | Leu ⇒ Pro | L483P |
CM057072 | - | AAC ⇒ AGC | 501 | Asn ⇒ Ser | N501S |
</figtable>
Mutation Analysis
In our analysis we looked closer to the amino acid properties and their changing characteristics by mutation. We analysed the structural difference between wild type (WT) and mutation. We also considered their secondary structure and distinguished between helix (H), sheet (E) and loop (C). We also took two different substitution matrices into account, BLOSUM62 and PAM250. Point Accepted Mutation matrix has only positiv integer values as scores and is not symmetric. The score reflects the probability of a amino acid to mutate into another. In contrast the BLOcks SUbstitution Matrix has also negativ integers and is symmetric. A positive score indicates that a substitution occurs more than random. While a score of 0 shows that the substitution occurs randomly, a negative one points to a mutation less frequent than a random mutation. In case one of our selected mutations has the worst possible substitution score for this amino acids we highlighted the score red in <xr id="ana"/>. To consider also evolutionary information we created different PSSM matrices. These position specific scoring matrices are based on alignments. Just as BLOSUM, the PSSM has positive and negative integer values as scores. A positve value shows that the substitution occurs more often than expected. Critical functional residues, like active site residues, have high positive scores. One PSSM was created with a PsiBlast search. The other one is basd on an alignment consisting of all mammalian homologous sequences.
Getting a bit closer to evolution you will have to create a PSSM (position specific scoring matrix) for your protein sequence using PSI-BLAST (5 iterations). How conserved are the WT residues in your mutant positions? How is the frequency of occurrence (conservation) for the mutant residue type? Anything interesting? And another step close to evolution: Identify all mammalian homologous sequences. Create a multiple sequence alignment for them with a method of your choice. Using this you can now calculate conservation for WT and mutant residues again. Compare this to the matrix- and PSSM-derived results. <figtable id="ana">
Mutation Analysis | |||||||
---|---|---|---|---|---|---|---|
Changes of Physiochemical Properties | Structural Properties | Conservation | |||||
Mutation | From | To | Pymol Visualization | Secondary Structure | BLOSUM62 score | PAM250 score | PSSM score |
S77R | polar, neutral charge, sulfur-containing | polar, positive, basic | E | -1 | 6 | 1 | |
N141S | polar, neutral charge, acidic | polar, neutral, sulfur-containing | H | 1 | 5 | 0 | |
R159Q | polar, positive charge, basic | polar, neutral, acidic | E | 1 | 5 | -4 | |
L213F | nonpolar, neutral charge, aliphatic, hydrophobic | nonpolar, neutral, aromatic, hydrophobic | E | 0 | 13 | 3 | |
G241E | nonpolar, neutral charge, aliphatic | polar, negative, acidic | C | -2 | 9 | -1 | |
V349I | nonpolar, neutral charge, aliphatic, hydrophobic | nonpolar, neutral, aliphatic, hydrophobic | E | 3 | 4 | 0 | |
T408M | polar, neutral charge, hydroxyl-containing | nonpolar, neutral, sulfur-containing | H | -1 | 5 | -1 | |
N409S | polar, neutral charge, acidic | polar, neutral, sulfur-containing | H | 1 | 5 | 1 | |
L483P | nonpolar, neutral charge, aliphatic, hydrophobic | nonpolar, neutral, cyclic | E | -3 | 5 | -3 | |
N501S | polar, neutral charge, acidic | polar, neutral, sulfur-containing | E | 1 | 5 | -2 |
</figtable>
Substitution and Phenotype
blossom
PSSM
Multiple Sequence Alignment
Comparison of different approaches
SIFT, Polyphen, Mutationtaster, SNAP -> one table
References
http://en.wikipedia.org/wiki/Amino_acid
[GENE%20AND%20%22human%22[ORGN]%20AND%20%22snp%22[SNP_CLASS]%20&cmd=DetailsSearch dbSNP]