Calculate the Molar Extinction Coefficient, and Weight/Volume Extinction Coeffic

Question

Calculate the Molar Extinction Coefficient, and Weight/Volume Extinction Coefficient (i.e., in g-1Lcm-1) for the following protein both under fully reduced and fully oxidized conditions using the equation to calculate extinction coefficients. The A280 of this protein sample made by diluting 50 L of the original stock to 1000 L under a fully reduced condition is 0.45. Calculate the concentration in mg / mL of the original protein stock that is under fully reduced condition. Assume a 1 cm path length. The molecular weight of the protein is 11716 Da.

Protein sequence:

MTAHILTAEAEAWLRGIIRYTRREWGAAQVRRYIAKLELCEMDYLAAGEGPFKWMSELYPALRMARCEHHYVRGADLPAGEPALVVAILHERMDLCTRLADRLKG

Explanation / Answer

Let the original stock solution concentration be = 5 mg/ml

concentration of final diluted solution = 5 x 0.05/1 = 0.25 mg/ml

concentration of protein in M = 0.25 x 1000/11716 x 1000 = 2.134 x 10^-5 M

Absorbance = ebc

e = molar extinction coefficient

e = 0.45/2.134 x 10^-5 x 1 = 21087.16 M-1.cm-1

   = 21087.16 x 11716

   = 2.47 x 10^8 L/g.cm-1

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