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Flashcards in Exam 2 Deck (146)
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1

What drives the protein folding? Why does it do?

hydrophobic effect

cause the non-polar groups to aggregate to minimize its contact with water

2

What does the protein folding to the energy and its entropy?

from high energy, high entropy to low energy, low entropy

3

What are the other cross-links (bonds) that help stabilize the protein after folding?

ion pairs (side chains -R between negative and positive)

disulfide bonds (Cys)

Zn2+

 

4

Is it true that protein adopt its 3rd comformation before its amino acid fully synthesized?

True

5

What can cause to denature protein? How does the factors do? 

Temperature, pH, detergent, chaotropic agents

disrupt the forces that hold protein in 3rd conformation

 

6

What happen to the misfolding protein? 

they can be sent back to get refold or degraded to its components amino acids.

7

What happen to misfolding protein that doesnt degrade or refold? What type of protein does this?

they aggregate 

Tau and amyloid-beta in Alzheimer's disease

prion protein in Cow's disease

alpha-synuclein in Parkinson

 

8

Why some proteins cant be degraded by protease?

because the fibers are resistant to degradation

9

What detergent can denature protein? What does mercaptoethanol do?

urea/ salt

cleave sulfide bonds in 3rd conformation.

10

What are 3 types of protein seperation? 

By charge

By binding specificity

By size

11

 Technique for seprate protein by charge

Technique for seprate protein by binding specificity

Technique for seprate protein by size

ion exchange chromatography

affinity chromatography

Gel filtration & SDS gel electrophoresis

12

In general, protein seperation requires what for every same technique?

Mobile phase (buffered solution with protein in it)
stationary phase

13

In ion exchange chromatography, what are the stationary phases? What does each do?

DEAE ( anion exchange: binding with negative group)

CM (cation exchange: bidning with positive group)

14

In ion exchange chromatography, how do we seperate the protein bound to stationary phase - DEAE?

Add high sal-solution or small pH solution

15

In ion exchange chromatography, how do we seperate protein bound with CM phase?

high salt-solution and higher pH

16

In what technique separating protein, are the PIs of amino acids important?

Ion exchange chromatography

17

What is the most efficient purification method? Why? 

affinity chromatography

Because the purification depends on the specific binding of the protein and the stationary phase (resin)

18

How do you seperate the protein bound to resin in affinity chromatography?

add high salt solution or ligands to compete binding btw protein and resin

19

What properties of proteins do the gel filtration chromatography and SDS-page use to purify proteins?

their size and shape

 

20

Which will move through column faster in gel filtration chromatography: larger or smaller protein?

larger proteins

21

What is the only technique that doesnt use high salt/ pH solution to purificate proteins?

gel filtration chromatography and SDS-page

22

Process of SDS-page purification

SDS detergents make all protein negatively charged

negative charges move toward positive end

bigger proteins move more slowly through the matrix

23

Sequencing steps for a protein

1) Purification of protein

2) The individual polypeptide chains are seperated

3) Large polypeptide are broken into smaller pieces (<100 residue)

4) Edman degradation

5) Piece together sequence from overlapping fragments 

24

What proteins are used to cleave peptide bonds? What is the characteristic of this protein? What is the process of this called?

protease (only cleave at the specific residues)

hydrolysis (adding water)

25

What is mass spectrometry used for?

analysis of amino acid sequences

26

Tandem MS-MS. How does this technique determine the amino acid?

first MS isolate a fragment, the second determine mass-to-charge ratios of fragment. 

By comparing all of fragments, compare those that different by mass of one amino acid to determine sequence.

27

What are techniques to analyze 3rd and 4th protein structure?

X-ray crystallography

cryoelectron microscopy

NMR

28

X-ray crystallography

 

Technique to determine the atomic and molecular structure

Crystallized protein is beamed with X-ray, leaving the X-ray diffraction pattern on X-ray film. This will be determined by computer to show 3D map of electron

29

What does the X-ray diffraction pattern show us? What does X-ray crystallography help in protein structure analysis?

position and intensities of spots on film

trace the polypeptide backbone and some general side chains

30

cryoelectron microscopy

What is this technique used for?

proteins structure are determined by using electron microscopy and then freezed (-196 celcius)
large/complexed proteins (ribosomes)