Week 9-10 Recitation Slides

Question 1

Why are proteins purified? [Practical Applications]

Answer 1

-clinical use (pharmaceutical agents),
-industrial uses (detergents or emulsifying agents) a
-reagents for research (restriction endonucleases, protein MW standards)

Question 2

Parameters to separate proteins

Answer 2

-physical (size, shape, charge, hydrophobicity, solubility) - biological properties (specific
interactions)……gel filtration and affinity chromatography

Question 3

Protein Purification Strategy (5 steps)

Answer 3

Extraction: Spheroplast
Clarification: centrifugation
Capture: concentrate by ammonium sulfate precipitation
Intermediate: Heat denaturation
Polishing: Ion-exchange chromatography

Question 4

Alkaline Phosphotase (properties, function, location in the cell, optimal conditions)

Answer 4

AP is a ubiquitous enzyme, Zn-containing enzyme, dimer, heat stable, MW 86,000.

Function is to remove phosphate groups from
phosphorylated compounds facilitating transport across membranes and providing the cell with a source of inorganic phosphate

located in the periplasmic space

maximum activity at high PH

Question 5

Why is E. Coli the bacteria of choice in biochemistry?

Answer 5

it is easy and rapid to grow in media, high transformability