Lecture 5 Membrane proteins

Question 1

Why do we study membrane proteins?

Answer 1

• 30% of sequenced genes encode membrane proteins• membrane proteins are involved in vital cellular processes (signal transduction, communication, transport)• 70% of drug targets are membrane proteins• many diseases due to compromised folding/processing of membrane proteins (Cystic Fibrosis, Alzheimer)• very few high resolution structures available

Question 2

Detergents are tools in MP research. What are their characteristics?

Answer 2

Amphiphilic, better soluble in water than lipids, charged or uncharged headgroup; strong detergents also denature proteins
•aggregate at higher conc. in micelles due to cone-shape, do not form bilayers
•CMC depends on temp, salt etc.

Question 3

What do detergents do? When is a detergent mild?

Answer 3

detergents disrupt lipid bilayer, hydrophobic tail binds to hydrophobic core of MP and brings it in solution through their very polar headgroups•in mild detergent, structure of MP is preserved

Question 4

How do you reconstitute functional membran proteins with the help of detergents?

Answer 4

•solubilization with mild detergent•purification•replace detergent with (natural) lipids•test membrane integration •test function•powerful tool to analyze function of MPs•powerful tool to determine the structure of MPs•powerful tool to analyze minimal requirements of particular membrane processes•not easy

Question 5

What do nanodiscs do?

Answer 5

Nanodiscs: new tool to solubilize MPs in active form

• dilute detergent and add lipids + HDL (high density lipoprotein)
• HDL forms belt round lipids ! soluble disc
• powerful tool for “single particle” analysis (structure and function)