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Designing this teaching tidbit has gotten completely out of hand. Group 1: Chemistry/Biology Interface Protein Folding. Xinnian Chen, U Conn. Alison Hill, Duke Ron Grunwald, Duke Dan Kiehart, Duke Dan Mulkey, U Conn. Carolyn Norris, Johns Hopkins Joel Schildbach, Johns Hopkins

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Designing this teaching tidbit has

gotten completely out of hand


Group 1: Chemistry/Biology Interface

Protein Folding

Xinnian Chen, U Conn.

Alison Hill, Duke

Ron Grunwald, Duke

Dan Kiehart, Duke

Dan Mulkey, U Conn.

Carolyn Norris, Johns Hopkins

Joel Schildbach, Johns Hopkins

Thanks to Amy Prunuske and Brad Hyman


  • Course: Introductory level course in which chemical foundations of biological function are described.

  • Students have been exposed to:

  • Basic chemistry of covalent and noncovalent bonds

  • DNA structure and sequence

  • Protein primary sequence is (almost always) specified by sequences in DNA.

  • mRNA is translated on ribosomes

  • Linear polypeptide chain, comes off the ribosome and begins to fold.


Thus they have been exposed to:

Text


UNIT: INTRODUCTION TO PROTEINS

Goal 1: ProteinPrimary Structure = Amino acid sequence.

Goal 2: ProteinSecondary, Tertiary and Quaternary Structure

Goal 3, TIDBIT: Explore HOW proteins fold


3.TIDBIT Goal: Students will begin to understand the mechanism by which proteins fold into higher order structures due to noncovalent interactions.

a. Outcome: Students will be able to evaluate experimental data to decide whether or not a protein can fold spontaneously.

b. Outcome:Students will be capable of predicting and describing the mechanism of disruption of protein structure by a chemical perturbant.

c. Outcome: Students will recognize that protein 3D structure is required for function.

(CN)

(JS)


Proteins with different

functions have different structures

  • Proteins with different shapes and sizes

  • Proteins reproducibly fold from their primary structures

  • Shape determines function

(JS)


Even small proteins have very complex 3D structures:

Ribonuclease: (RNase) is a digestive enzyme

cuts RNA polymers into monomers

124 amino acids

>900 atoms (not counting hydrogens!)


Three different 3D representations of ribonuclease:

Ribbon Diagram Highlights Secondary Structure

Stick Diagram Shows the Position of Heavy Atoms (not H)

Space Filling Model Features Surface Topology


Hydrogen bonds stabilize protein structure, e.g., alpha helices

H-bond

(JS)

(DM)


Urea offers an opportunity to investigate the bonds that stabilize protein structure: How?

(DM)


How does urea interact with proteins?1) 1 minute drawing: show the interactions of 6 molecules of Urea with the polypeptide backbone on the handout

(DM)


2) Think/Pair/Share: What happens when

you add 8M Urea (a REALLY HIGH CONCENTRATION) to 1 µM alpha helix?

Collectively choose a hypothesis for what 8M Urea does to a protein...

(DPK)

(DM)


Next, we’ll use Urea as a probe of protein structure in an experiment

(DPK)


Experimental Data

Ribonuclease

in

salt solution

+/- Urea

?

(DPK)


Clicker Question:

What do you think is the most likely effect of Urea on the STRUCTURE of the RNase?

A

B

C

Ribonuclease

in

dilute salt water

+ 8M Urea

D

(DPK)

0%

activity

100%

activity

100%

activity

Ribonuclease,

dilute salt water,

Urea removed


A

B

C

Ribonuclease

in

dilute salt water

+ 8M Urea

D

Clicker Vote

Table discussion: rationale behind your choice

(Revote if wide distribution)

Class summarize why?

(DPK)

(RG)


Cut into Pieces? Aggregate? Specific Inhibitor?

You’ll learn later that good inhibitors function at nM concentrations:

Remember that Urea works at M concentrations!

(DPK)

(RG)


Christian Anfinsen:

Nobel Prize in Chemistry - 1972

  • One minute essay:

  • Based on what you’ve learned, write a that relates structure, function and reversible folding/unfolding

(RG)


  • For next class: Develop a hypothesis of protein folding that relates primary sequence to native structure and function

(XC)


Adios!


?

Ribonuclease,

dilute salt water,

+ “Stuff”

Ribonuclease,

dilute salt water,

“Stuff” removed

Ribonuclease

in

dilute salt water


  • the “Stuff” is Urea (cute huh?):

  • Clicker Question: What kind of bond/force is Urea most likely to disrupt?

  • A. covalent B. ionic C. van der Waals D. hydrogen E. traxoline


  • Adios!


  • Catalog and know NC/sidechain interaction

  • Assessment LOCS

  • Manipulate strings, evaluate

  • Specific inteactions

  • Unique Structures

  • Next anfinsen with beta mercaptoethanol?


what kind of interactions mediate these different levels of structure?

Focus on noncovalent bonds and the SS bonds that stabilize it


Secondary Structure: H-Bonds between atoms in the Polypeptide Backbone


Structure of alpha helix and urea

what happens and how?


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5 possible

partners

7 possible

partners

3 possible

partners

just one

option left

3 possible

partners


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