Computation of various physical and chemical properties of Proteins and DNA from their primary sequence

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Computation of various physical and chemical properties of Proteins and DNA from their primary sequence

Procedure

Protein

  1. Approximate molecular weight of protein

    Approximate molecular weight of protein = Number of amino acids x 110 Da (molar mass: 1g/mol = 1 Da)

    Amino Acid 1-letter code Molecular weight (g/mol)
    Alanine A 89.1
    Arginine R 174.2
    Asparagine N 132.1
    Aspartate D 133.1
    Cysteine C 121.2
    Glutamate E 147.1
    Glutamine Q 146.2
    Glycine G 75.1
    Histidine H 155.2
    Isoleucine I 131.2
    Leucine L 131.2
    Lysine K 146.2
    Methionine M 149.2
    Phenylalanine F 165.2
    Proline P 115.1
    Serine S 105.1
    Threonine T 119.1
    Tryptophan W 204.2
    Tyrosine Y 181.2
    Valine V 117.1
  2. Actual molecular weight of a protein

    In order to calculate actual molecular weight of a protein, the following step must be followed
    Suppose there is a protein with the amino acid sequence MRANLDFW

    This protein has 7 peptide bonds, and during formation of each peptide bond 1 molecule of water having molecular mass of 18 g/mol is removed. So molecular mass of this protein would be

    Molecular weight of (methionine(M)+Arginine(R)+Alanine(A)+Asparagine(N)+Leucine(L)+Aspartate(D)+Phenyalanine(F)+Trytophan(W) – (7x18)) [ Molecular weight of water-18 gm]

  3. The molar absorption coefficient of a peptide or protein

    is related to its tryptophan (W), tyrosine (Y) and cysteine (C) amino acid composition. At 280nm, this value is approximated by the weighted sum of the 280nm molar absorption coefficients (ε) of these three constituent amino acids, as described in the following equation:

    ε = (nW×5500) + (nY×1490) + (nC×125)

    where n is the number of each residue and the stated values are the amino acid molar absorptivities at 280nm.

    Molar absorptivities have units of M-1 cm-1.

  4. Isoelectric pH(pI) calculation
    • For amino acids with neutral sidechains, the pI can be calculated by averaging the pKa values of the carboxylic acid and ammonium groups
    • For amino acids with acidic sidechains, the pI can be calculated by averaging the pKa values of the two most acidic groups
    • For amino acids with basic sidechains, the pI can be calculated by averaging the pKa values of the least acidic groups
  5. For calculation of pI of a protein

    Step 1: We must first write the pKa values of the amino acid constituting the protein from low to high.
    Step 2: Select a pH value below the lowest pKa value of the amino acid and determine the net charge of the amino acid at that pH.
    Step 3: Select a pH between the first and the second pKa values of the amino acid and determine the net charge of the amino acid.
    Step 4: Select a pH between the second and the third pKa value of the amino acid and determine the net charge of the amino acid.
    Step 5: Raise the pH above the highest pKa value and determine the net charge of the amino acid.
    Step 6: Calculate the pI by averaging the two pKa values that are just before and just after the zero net charge.

DNA

  1. The average weight of a single DNA bp is 660 daltons. This can also be written as 660 g/mol (molar mass: 1g/mol = 1 Da). The molecular weight or molar mass of any double-stranded DNA fragment can therefore be calculated by multiplying number of nucleotides by 660 and the answer will be expressed as daltons or g/mol.
  2. Actual molecular mass of double stranded DNA
    Formula for calculating actual molecular mass of double stranded DNA
    M.W. = (nA x 313.2) + (nT x 304.2) + (nC x 289.2) + (nG x 329.2)
  3. Melting temperature of DNA is calculated by the formula
    Tm = (nA+nT) * 2 + (nG+nC) * 4 °C