protein folding, madison 2009

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Dec 14, 2013 (3 years and 10 months ago)

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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.


Text

Thus they have been exposed to:

UNIT: INTRODUCTION TO PROTEINS


Goal 1: Protein

Primary Structure = Amino acid sequence.


Goal 2:

Protein

Secondary, 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)




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




(DM)

(DPK)

2) Think/Pair/Share: What happens when

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

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?


Ribonuclease,

dilute salt water,

Urea removed

D

C

A

B

Ribonuclease

in

dilute salt water

+ 8M Urea

(DPK)

100%

activity

0%

activity

100%

activity

Table discussion: rationale behind your choice


(Revote if wide distribution)


Class summarize why?

Clicker Vote

D

C

A

B

Ribonucleas
e

in

dilute salt
water

+ 8M
Urea

(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)

One minute essay:


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

Christian Anfinsen:

Nobel Prize in Chemistry
-

1972


(RG)

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

(XC)

Adios!

Ribonuclease

in

dilute salt water

Ribonuclease,

dilute salt water,

+ “Stuff”

Ribonuclease,

dilute salt water,

“Stuff” removed

?

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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7 possible

partners

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partners

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3 possible

partners

3 possible

partners

just one

option left