Question

You are asked to purify a hexokinase. Discuss the parameters you would consider in sourcing and selecting such an enzyme, and briefly indicate how you would stabilize these proteins on isolation. In answering this question, you should ask and answer the following questions.

A. What is a hexokinase?

B Which hexokinase would you think of purifying? Why? Think where would you find lots of this enzyme in a mammal? (You should be able to get this organ easily at the butcher’s from a pig or cow.)

C will you find this enzyme in highest concentration in liver or muscle? Why?

D Give details of how you will extract, isolate and purify the enzyme.

E How would you prove that you have indeed extracted a/this hexokinase?

Answer 1

1. Hexokinase is an enzyme which phosphorylate the hexoses by transfer of phosphate from ATP to hexose.

2. Most common type of hexokinase is glucokinase which syntgesize Glucose-6-Phosphate from Glucose. This enzyme is easily extracted from human heart and 1 kg of human heart tissue contain 25 mg of hexokinase.

3. Hexokinase is present in liver in hepatocytes in highest conc. and control glycogen formation and breakdown and sugar uptake.

4. Glucokinase is in higher concentration in heart tissue in humans and 1 kg of heart tissue give rise to 25 mg of hexokinase and which can be purified by using affinity chromatography by Glucosamine.

6. Glucokinase can be characterized by using Glucose-6-Phosphate for dimerisation of glucokinase.