You are presented with the following data from a series of purification steps fo

ID: 167854 • Letter: Y

Question

You are presented with the following data from a series of purification steps for the enzyme comatose. Fill in the Blanks Below Given the purification schemed data show in problem 2. which purification step gave the greatest: Relative increase in product purity (the greatest Increase from previous step)? loss of product? Absolute reduction of product (i.e., the greatest decrease in actual volume)? Related reduction of product volume (the greatest% decrease from the previous step)? Suppose you are given the assignment of isolating an enzyme, "enslave" which is secreted in to the fermentation broth by the bacterium a. the enzyme has an approximate molecular weight of 60.000. You grow 3L of B. which you then spin down in a centrifuge. You discard the broth and weight the cells. You break open the Celts with a grinding method and centrifuge the resultant paste. You discard the cellular debris

Explanation / Answer

1. FORMULAS FOR CALCULATION

VOLUME

TOTAL PROTEIN

TOTAL ACTIVITY

SPECIFIC ACTIVITY

YIELD (%)

PURIFICATION FACTOR

HOMOGENIZATION

10000

12350

9000

0.73

100

DIALYSIS

5000

10233

8289

0.81

92.1

1.11

ORGANIC EXTRACTION

3860

6369

1.65

70.76

2.26

ION EXCHANGE CHROMATOGRAPHY

1140

5625

4.93

62.5

6.75

PAGE

386

4688

12.15

52.09

16.64

2. a)RELATIVE INCREASE IN PRODUCT PURITY (GREATEST INCREASE FROM PREVIOUS STEP): From the table by comparing the PURIFICATION FACTOR we can see that in PAGE there is the greatest increase in product purity from previous step.

b)LOSS OF PRODUCT: From the table by comparing the yield percent we can say that except homogenization all the purification step viz DIALYSIS, ORGANIC EXTRACTION, ION EXCHANGE CHROMATOGRAPHY & PAGE leads to product loss.

c)ABSOLUTE REDUCTION OF PRODUCT (GREATEST DECREASE IN ACTUAL VOLUMN); From the table by comparing the yield (%) column we can observe that that yield percent in PAGE is least i.e. 52.09 % which indicates that in case of PAGE there is a greatest decrease in actual volume.

d)RELATIVE REDUCTION OF PRODUCT (GREATEST % DECREASE FROM PREVIOUS STEP): From the table by comparing the yield (%) column we can observe that that GREATEST % DECREASE OF PRODUCT FROM PREVIOUS STEP occurs in ORGANIC EXTRACTION step