PAH-gene-Versus-PAH_allele(R408W)


A narrative blog post which includes an original description of the allele and a discussion of the impact of the change on the predicted secondary structure.

0.0001 PHENYLKETONURIA
PAH, IVS12DS, G-A, +1
The first PKU mutation identified in the PAH gene was a single base change (GT-to-AT) in the canonical 5-prime splice donor site of intron 12.

Since the variant/allele 0.0001 is due to the single base change in the site of intron 12, I picked the allele 0.0002 and discussed the impact on the predicted secondary structure.

0.0002 PHENYLKETONURIA
PAH, ARG408TRP
This defect is caused by a CGG-to-TGG transition in exon 12, resulting in an amino acid substitution (arg-to-trp) at residue 408 (R408W) of PAH.
This mutation makes up about 20% of the mutant PAH genes. It is one of the examples of CpG mutation. The R408W mutation occurs within the catalytic domain of PAH. PAH with the R408W mutation formed high-molecular-mass aggregates, indicative of severe distortion of the protein's oligomeric state.

I opened the mRNA sequence of my PAH gene in the seq-builder and highlighted the open reading frame of the gene beginning with the ATG codon of methionine. I highlighted the ORF in the upper strand and translated it into the protein sequence and saved it.
Since the allelic/variant is the change in the amino acid Arginine to(R) to tryptophan (W), I opened the protein sequence in the edit seq made the change and saved it under the name of PAH-allele(R408W).  
The Arginine is a basic amino acid and tryptophan is a hydrophobic amino acid thus I expect to see changes in the secondary structure. 

The protein statistics in the edit-seq about the PAH-gene and the PAH-allele is different in the molecular weight, isoelectric point to PH.
The PAH-gene Protein statistics.
    Molecular Weight 51862.15 Daltons 
      452 Amino Acids
    54 Strongly Basic(+) Amino Acids (K,R)
        60 Strongly Acidic(-) Amino Acids (D,E)
        155 Hydrophobic Amino Acids (A,I,L,F,W,V)
        119 Polar Amino Acids (N,C,Q,S,T,Y)
        6.408 Isolectric Point
        -4.147 Charge at PH 7.
    Protein Info about pah-allele-R-W.pro(1,453)
           Molecular Weight 51892.17 Daltons
         452 Amino Acids
       53 Strongly Basic(+) Amino Acids (K,R)
          60 Strongly Acidic(-) Amino Acids (D,E)
         119 Polar Amino Acids (N,C,Q,S,T,Y)
        6.293 Isolectric Point 
     -5.147 Charge at PH 7.0 
The changes seen in the Protean of DNASTAR are as following:
The Beta Regions from the amino acid 400 till the end of the secondary structure as depicted by the Garnier-Robson and Chou- Fasman algorithm.  The turn regions by the Chou-Fasman algorithm. The coil region by the Garnier-Robson algorithm the alpha amphipathic region and the Beta Amphipathic region as well as the flexible regions by Karplus-Schulz algorithm.
I have observed changes in the Apha, beta coil and turn regions by the Chou-Fasman and Garnier-Robonson as seen in the zoomed version of the Protean in DNASTAR.
The PAH-gene-&&-PAH-allele(R408W)


The detalied comparison of the PAH gene and its allele with (R408W) in Protean.




The changes seen in the region from amino acid 400 to the end of the polymer for the PAH gene and the allele variant in Protean. The Amino Acids from 400 to 420 are highlighted.
PAH-gene-400-420AA


PAH-allele(R408W)-400-420AA



The secondary structure changes as seen in the Protean 3D of DNASTAR in the PAH-gene and PAH-alllele (R408W).
Secondary Structure - Garnier-Robson—Examines the propensity of a given residue to exist in a certain structure.
I observed the changed in the beta regions, turn regions and coil regions in PAH-gene and allele but no change in the alpha regions.


PAH-gene-400-420AA-highlighted
PAH-allele(R408W)-400-420AA-highlighted
.
Secondary Structure - Chou-Fasman—Predicts secondary structure of proteins from the crystallographic structures of their mino acid sequences.
 I did not observe any change in the secondary structure PAH-gene and its allele.

PAH-gene-400-420AA-highlighted

PAH-allele(R408W)-400-420AA-highlighted


Hydropathy - Kyte-Doolittle—Predicts regional hydropathy of proteins from their amino acid sequences.
The hydrophicity and the hydrophobic regions were different in the PAH_gene and its allele (R408W) as expected. No change was noticed in the hydrophilic regions.

PAH-gene-400-420AA-highlighted

PAH-allele(R408W)-400-420AA-highlighted


Hydropathy - Hopp-Woods—Finds protein antigenic determinants by searching protein sequences for the area of greatest local hydrophilicity.
The hydrophicity and the hydrophobic regions were different in the PAH_gene and its allele (R408W) as expected. No change was noticed in the hydrophilic regions.

PAH-gene-400-420AA-highlighted
PAH-allele(R408W)-400-420AA-highlighted



Antigenicity- Jameson-Wolf—Predicts potential antigenic determinants by combining existing methods for protein structural predictions.
The antigenicity regions of the two structure are also different.


PAH-gene-400-420AA-highlighted

PAH-allele(R408W)-400-420AA-highlighted

Amphiphilicity – Eisenberg—Predicts the Eisenberg Moment.

Due to the incorporation of the hydrophobic amino acid tryptophan in the allelic variant the Amphilicity of the PAH-gene and its allele was different in the Hydrophobicity plot, Alpha amphipathic and Beta amphipathic region and Alpha and Beta moment.
PAH-gene-400-420AA-highlighted
PAH-allele(R408W)-400-420AA-highlighted

Surface Probability – Emini—Predicts the probability that a given region lies on the surface of a protein. The surface probability for the region between the amino acid 400 to 420 for the PAH-gene and PAH-allele (R408W) was also changed.
PAH-gene-400-420AA-highlighted
PAH-allele(R408W)-400-420AA-highlighted

Flexibility - Karplus-Schulz—Predicts backbone chain flexibility.
The PAH-gene and its mutant had different flexibility plots in the region of Amino Acids 400 to 420.
PAH-gene


PAH-allele(R408W)

Even the stability of the two structure showed difference in their plots in the amino acid 400-420 region.
PAH-gene

PAH-allele

Thus I observed marked differences in  the secondary structure of  the PAH gene and its allele (R408W) in the Amino Acid regions from 400 to 420 amino acids. 

No change was observed in the transmembrane property between the PAH-gene and its allele.
PAH-gene

PAH-allele



The vocabulary used in the description of the gene and allele
Linkage disequilibrium
The nonrandom association between two or more alleles such that certain combinations of alleles are more likely to occur together on a chromosome than other combinations of alleles
In other words, linkage disequilibrium is the occurrence of some combinations of alleles or genetic markers in a population more often or less often than would be expected from a random formation of haplotypes from alleles based on their frequencies.
Haplotype
A haplotype is a set of DNA variations, or polymorphisms, that tend to be inherited together. A haplotype can refer to a combination of alleles or to a set of single nucleotide polymorphisms (SNPs) found on the same chromosome.
Oligomer
A polymer that consists of two, three, or four monomers.
A monomer may combine with another monomer through chemical bonds to form a larger molecule, as in a polymer.
Examples of monomers are amino acids that link together by a peptide bond forming a polypeptide or protein.







No comments:

Post a Comment