A biochemist discovers and purifies a new enzyme, generating...

A biochemist discovers and purifies a new enzyme, generating the purification table below. Purification Step | Total Protein (mg) | Activity (units) 1. Crude Extract | 20,000 | 4,000,000 2. Precipitation (salt) | 5,000 | 3,000,000 3. Precipitation (pH) | 4,000 | 1,000,000 4. Ion-exchange chromatography | 200 | 800,000 5. Affinity chromatography | 50 | 750,000 6. Size-exclusion chromatography | 45 | 675,000 a. From the information in the table, calculate the specific activity of the enzyme after each purification step. b. Which of the purification steps used for the enzyme is the most effective (i.e., gives the greatest relative increase in purity)? c. Which of the steps is least effective? d. Is there any indication based on the results shown in the table that the enzyme after step 6 is now pure? e. What else could be done to estimate the effectiveness of the purification procedure, the purity of the final sample, and estimate the molecular weight of the enzyme?

Transcript

00:01 Hello everyone.

00:02 In this video we are going to calculate the specific activity of the enzyme after each purification procedure.

00:09 In the question, different purification steps are given along with total protein and activity of the enzyme.

00:17 The formula of specific activity to calculate is total activity of the enzyme divided by total protein.

00:24 So the first step is the crop extract.

00:27 This is the total activity and total protein b1.

00:29 Dividing this value will get specific activity for the state which is 200 micromole per m g minute the second step is precipitation with salt this is the total activity dividing it with total protein will get specific activity which is 600 micromole for mg minute this is the start step which is precipitation with ph this is the total activity dividing it with total protein will get specific activity which is 250 micro mole per ng minute and then this is the fourth step iron exchange chromatography this is the total activity and the total protein given dividing this value will give specific activity which is 4 ,000 micrmode per m g minute and then fifth step affinity chromatography here this is the total activity dividing it with total protein will get specific activity which is 15 000 microme per m g minute and the last step is six step, which is size exclusion chromatography.

01:34 This is the total activity.

01:36 Dividing it with total protein will get specific activity, which is 15 ,000 microman per ang permanency.

01:43 So here, specific activity has been increased with the successive steps of the purification of the enzyme.

01:52 The next part of the question we have asked, which of this procedure is most effective? so to understand this we have to look for the specific activity value.

02:03 So we can see that the specific activity value has been increased.

02:07 But here the specific activity gives the measurement of the purity of the enzyme in the mixture and it will increase as the purification progresses.

02:20 And further, the specific activity of the enzyme will be stable and same as the presentation reciting step where we can say that the octane enzyme is highly pure.

02:36 So here we can see that the specific activity of the sixth step is highest.

02:42 So the answer is size exclusion chromatography is most effective.

02:49 Then we have to estimate which of the procedure is less effective...

Question

Please give Ace some feedback