Biochemistry Test 2: Amino Acids And Protein Analysis

Front 1

3 Analytical identification by charge/mass

Back 1

1. Electrophoresis
2. Isoelectric focusing
3. 2D gel electrophoresis

Front 2

Column chromatography purification

Back 2

By ion exchange, size exclusion, or affinity

Front 3

Electrophoresis

Back 3

Mobility = total charge/friction= distance
Friction = mass
Direction of movement depends on the charge

Front 4

SDS gel electrophoresis(PAGE)

Back 4

1. Remove the charge so only mass counts
2. denature protein
3. encapsulate in SDS micelles to give a uniform charge
4. gives protein size
5. Compare mobility to a standard to get estimated mass

Front 5

Isoelectric focusing

Back 5

Using a pH gradient
1. Put ampholytes into one end of the gel
2. Electrophores them and let them migrate to their respective pHs
3. Put in your protein and electrophores them
4. They will migrate to their isoelectric point at their pH

Front 6

2D gel electrophoresis

Back 6

Seperating the molecules first by charge and then by mass with SDS.
This allows us to find abnormal proteins in disease states

Front 7

Proteomics

Back 7

trying to identify all proteins in an organism

Front 8

Metabolomics

Back 8

Trying to identify all metabolites in an organism

Front 9

Acute phase response

Back 9

General response of the body to injury or infection of tissue.
Production of some plasma proteins increase like; C reactive protein, protease inhibitors(antitrypsin), blood coagulation proteins, alpha 1 and 2 globulins
Other proteins decrease: albumin, transretin

Front 10

CRP(C reactive protein) as a marker?

Back 10

A marker because it is a major component of acute phase response to bacterial infection

Front 11

Affinity chromatography

Back 11

Having a specific ligand for the target protein then later disrupting that bond to purify it

Front 12

Size exclusion

Back 12

Larger ones pass through more quickly because the need to slip through the cracks
The smaller ones can take their time and also diffuse through the porous proteins making it easier and slower

Front 13

UV absorbance

Back 13

Directly proportional to the concentration of Phe, Tyr and Trp at 260-280nm.
The T's are more absorbant than P

Front 14

DNFB and Dansyl chloride

Back 14

Dinitroflourobenzene gives flourescent N terminals and detects picomolar amounts

Front 15

Ninhydrin reaction used as a detector?

Back 15

Detects nanomolar amounts

Front 16

Edman(endo) degradation

Back 16

Breaks down proteins in repetitive cycles using endonucleases. They are then seperated by chromatography

Front 17

Proteolytic enzymes and targets, chart of of the enzymes targeting AA bonds?

Back 17

1. Trypsin - Lys, Arg
2. Chymotrypsin - Tyr, Phe, 3.Ile, Leu, Val, Trp, His
4.Pepsin - Phe, Leu
5.Thrombin - Arg
6.Papain - Arg, Lys, Phe
7.Carboxypeptidase - C terminal residue
8. Use endoproteases to sequence proteins