A9 Isolation and Purification of Proteins

Question 1

ISOLATION OF CASEIN: Products (5)

Answer 1

MILK
(40C, 10% HCl, Centrifuge)

Curd
(Wash with distilled H2O until Cl- free filter)

Ppt.
(dry between filter paper, wash with 95% alcohol filter)

Ppt.
Ether
filter

Ppt.
Casein

Question 2

ISOLATION OF CASEIN: Discard

Answer 2

Whey

Filtrate

Filtrate

Filtrate

Question 3

ISOLATION OF EGG ALBUMIN: Products (4)

Answer 3

EGG WHITE
(1N HOAc
filter through cheesecloth, stir)

filtrate
(saturated (NH4)2SO4
centrifuge

Centrifugate
(saturated (NH4)2SO4
refrigerate 2-5 days
centrifuge

ppt albumin

Question 4

ISOLATION OF EGG ALBUMIN: discard

Answer 4

ppt Ovoglubin & ovomucin

Question 5

Removal of proteins from the chemical mass or biological fluid in which they are found through disruption of tissue

Answer 5

Isolation

Question 6

Purity is increased for every step of isolation performed on a protein

Answer 6

Purification

Question 7

Conditions controlled not to denature protein (3)

Answer 7

Low temperature

pH near protein native pH

High Protein concentration

Question 8

Procedures used for separation and purification of proteins (3)

Answer 8

Salting out

Isoelectric precipitations

precipitation with organic solvents or acidic protein precipitants

Question 9

Methods used to establish the purity or homogenity of protein preparation (4)

Answer 9

Testing solubility in a solvent

Electrophilic mitigation rate

Rate of sedimentation

Chromatographic analysis

Question 10

Chemicals which can be used to extract proteins (4)

Answer 10

Water

Dilute salt

Acid

Alkaline solutions

Question 11

Detergent solutions or organic solvents needed when protein is associated with other organic substances thus cannot be extracted

Answer 11

Acetone

Ether

Butanol

Question 12

Used when necessary to remove small ions or exchange one ionic environment (3)

Answer 12

Dialysis

Ultrafiltration

Moelcular exclusion chromatography