9780521516358book.pdf

Table 8.3

Example of a protein purification schedule

Fraction

Volume(cm

3 )

Proteinconcentration(mg U cm

^3

)

Totalprotein(mg)

Activity

a

(mg U cm

^3

)

Totalactivity (U)

Specificactivity(U mg

^1

)

Purificationfactor

b

Overallyield

c(%)

Homogenate

8 500

40

340 000

1.8

15 300

0.045

1

100

45%–70%(NH

) 42

SO

4

530

194

103 000

23.3

12 350

0.12

2.7

81

CM-cellulose

420

19.5

8 190

25

10 500

1.28

28.4

69

Affinity chromotography

48

2.2

105.6

198

9 500

88.4

1 964

62

DEAE-Sepharose

12

2.3

27.6

633

7 600

275

6 110

50

Notes:

aThe unit of enzyme activity (U) is defined as that amount which produces 1

mmole of product per minute under standard assay conditions.

bDefined as: purification factor

¼

(specific activity of fraction/specific activity of homogenate).

cDefined as: overall yield

¼

(total activity of fraction/total activity of homogenate).

Reproduced with permission from

Methods in Molecular Biology

,^59

,Protein Purification Protocols

, ed. S. Doonan (1996), Humana Press Inc., Totowa, NJ.