Calculating Enzyme Specific Activity
The specific activity of an enzyme is a measure of its catalytic efficiency per unit of protein. It is calculated using the formula:
$ \text{Specific Activity} = \frac{\text{Enzyme Activity}}{\text{Protein Concentration}} $
Step 1: Determine Enzyme Activity per Milliliter
The enzyme preparation has an activity of 2 Units per 20 $\mu$l. To find the activity per milliliter (ml), we first convert $\mu$l to ml:
- $20 \ \mu\text{l} = \frac{20}{1000} \text{ ml} = 0.02 \text{ ml}$
Now, calculate the activity in Units per ml:
$ \text{Activity per ml} = \frac{2 \text{ Units}}{0.02 \text{ ml}} = 100 \text{ Units/ml} $
Step 2: Identify Protein Concentration
The protein concentration is given directly:
$ \text{Protein Concentration} = 0.4 \text{ mg/ml} $
Step 3: Calculate Specific Activity
Using the formula for specific activity:
$ \text{Specific Activity} = \frac{100 \text{ Units/ml}}{0.4 \text{ mg/ml}} $
$ \text{Specific Activity} = \frac{100}{0.4} \ \frac{\text{Units}}{\text{mg}} $
$ \text{Specific Activity} = 250 \ \frac{\text{Units}}{\text{mg}} $
Therefore, the specific activity of the enzyme is 250 Units/mg.
Important Questions from Enzyme Assays Molar Extinction Coefficient
- A solution shows a transmittance of 20% when taken in a cuvette of 2.5 cm path length. If the molar absorption coefficient of the solution is $12000 \text{ dm}^3/\text{mol.cm}$, the concentration of the solution is ________ $\times 10^5 \text{ mol/dm}^3$ (rounded off to two decimal places).
- A solution containing GTP has molar extinction coefficient of $1.55 \times 10^4$ $mol^{-1}dm^3cm^{-1}$ at a given wavelength. The concentration of GTP solution is $1.290 \times 10^{-5}$ $mol$ $dm^{-3}$. The absorbance of GTP solution in 1 cm cuvette at the same wavelength will be .................
- Measurement of the absorbance of a solution containing NADH in a path length of 1cm cuvette at 340 nm shows the value of 0.31. The molar extinction coefficient of NADH is $6200 M^{-1} cm^{-1}$. The concentration of NADH in the solution is ________ $\mu M$ (correct to integer number).
- If a $10$ mM solution of a biomolecule in a cuvette of path length $10$ mm absorbs $90\%$ of the incident light at $280$ nm, the molar extinction coefficient of the biomolecule at this wavelength is ________ $M^{-1}cm^{-1}$. (Round off to two decimal places)
- The absorbance of a $5 \times 10^{-4}$ $M$ solution of tyrosine at 280 $nm$ wavelength is 0.75. The path length of the cuvette is 1 $cm$. The molar absorption coefficient at the given wavelength in $M^{-1}cm^{-1}$, correct to the nearest integer, is ________.
