Non-enzymatic Protein Function and Protein Analysis

Question 1

What is a motif?

Answer 1

A motif is when two secondary structures repetitively organize together.

• Ex: Two Beta sheets interacting

Question 2

What is collagen?

Answer 2

Collagen is a strong, yet flexible, extracellular structural protein comprised of three intertwined alpha helix motifs. It’s primary function is support of tissues outside the cell.

• Collagen makes up most of the extracellular matrix of the connective tissue.

Question 3

What is elastin?

Answer 3

Elastin is another structural protein found in the extracellular matrix of connective tissue.

• Can stretch and recoil - restores the shape of tissue

Question 4

What is Keratin?

Answer 4

Keratin makes up the intermediate filaments of epithelial cells.

• Keratin is essentially the cellular (cell’s) skeleton.
  
  • Give support and shape to each individual cell.

Question 5

What is actin?

Answer 5

Actin is a structural protein that makes up microfilaments and myofibrils.

• Major component of muscles
• It is the most abundant structural protein in eukaryotic cells.
• Actin is polar (has a positive and negative end)
  
  • Charge indicated by direction of movement

Question 6

What is tubilin?

Answer 6

Tubilin is a structural protein that aids in cell division (mitosis and meiosis) but is also the framework for intracellular transport between kinesin and dynein.

• Tubulin like actin is polar.
  
  • Positive end is away from the nucleus
  • Negative end is towards the nucleus

Question 7

Which structural proteins serve as the structural framework in which motor proteins can move?

Answer 7

Actin and Tubulin

Question 8

What binds together to form the thick filaments in a sarcomere?

Answer 8

Myosin fibers bind together to form the thick filaments in a sarcomere.

• By hydrolyzing ATP, the myosin heads can move along the actin myofibril causing the muscle to contract.

Question 9

What are kinesin and dynein responsible for?

Answer 9

Kinesin and dynein transport things within a cell (intracellular transport).

• Contain two ATPase domains that allow Kinesin and Dynein to rotate and walk down a microtubule.
• Kinesin and dynein move in opposite directions along Beta-subunits of tubulin.
  
  • Dynein towards nucleus (Dyne-in)
  • Kinesin away from nucleus (Karry-out)

Question 10

What are binding proteins?

Answer 10

Binding proteins are when subunits combine to form quaternary structure and transport molecules throughout the body.

Ex: Hemoglobin

Question 11

What are Cadherins?

Answer 11

Cadherins are cellular adhesion molecules (proteins) that adjoin cells that remain neighbors (stationary). They are not used for free-floating cells that need to temporarily stick to another cell.

• They participate in the calcium-dependent adhering of cells.
• The typically are found in intracellular junctions holding cells together.
  
  • Cadherins attach to other cadherins

Question 12

What are integrins?

Answer 12

Integrins are cellular adhesion molecules that are integrated into the cell membrane of one cell and bind to the extracellular matrix or another cell.

Question 13

What are selectins?

Answer 13

Selectins are cellular adhesion proteins that seclectively bind to carbohydrates on another cell.

Question 14

Selectins and integrins regulate what process involving neutrophils?

Answer 14

Selectins and integrins regulate neutrophil extravasion, which is when neutrophils exit the bloodstream and enter the surrounding tissue.

Question 15

What are immunuglobulins?

Answer 15

Immunoglobulins are antibodies. They are Y shaped proteins composed of two light chain proteins bonded to two heavy chain proteins via disulfide bonds (Cysteine). A disulfide bond is also formed between the two heavy chains.

Question 16

Where is the variable and constant region on an immunoglobulin (antibody)?

Answer 16

Question 17

What are 3 common types of ion channels?

Answer 17

1. Ungated Ion Channels (Diffusion)
2. Voltage-Gated Ion Channels
3. Ligand-Gated Ion Channels (most specific of 3)

Question 18

Which type of signalling receptor requires ATP to function?

Answer 18

G Protein-Coupled Receptors (GCPRs) rely on ATP because Adenylate Cyclase uses ATP to produce the signalling molecule cyclic AMP (cAMP).

Question 19

What are the different types of G Protein-Coupled Receptors in Eukaryotes?

Answer 19

Question 20

True or false: Motor proteins are not enzymes?

Answer 20

False: Motor proteins do not predominantly function as enzymes. However, they do display catalytic activity acting as ATPases.

• Enzymes are proteins or RNA molecules that display catalytic activity

Question 21

How do cytoskeletal proteins differ from motor proteins?

Answer 21

Cytoskeletal proteins tend to be fibrous with repeating domains (motifs), while motor proteins tend to have ATPase activity and binding heads.

• Actin and tubulin (cytoskeletal proteins) have polarity.
• Kinesin moves towards postivive end (periphery of the cell). Dynein moves towards negative end (nucleus)

Question 22

What could permit a binding protein involved in sequestration to havve a low affinity for its substrate and still have a high percentage of substrate bound?

Answer 22

If the binding protein is present in sufficiently high quantities relative to the substrate, nearly all substrate wil be bound despite a low binding affinity.

• High binding protein concentration with low substrate concentration.

Question 23

What are the 3 types of Cell Adhesion Molecules (CAMs) and what type of adhesion doeas each class form?

Answer 23

1. Cadherins
   
   • Cadherins bind to cells of the same or similar type using calcium (calcium-dependent binding)
2. Integrins
   
   • One cell to proteins in the extracellular matrix
3. Selectins
   
   • One cell to carbohydrates, usually on the surface of other cells

Question 24

What type of CAM is has alpha and beta chains?

Answer 24

Integrins have alpha and beta chains in the integral protein that are involved in binding one cell to the proteins in the extracellular matrix.

• Integrins are also involved in apoptosis and cellular division

Question 25

What type of CAM is expressed on white blood cells and the endothelial cells that line blood vessels?

Answer 25

Selectins are expressed on the surfaces of white blood cells and endothelial cells that line blood vessels.

• Selectins bind to carbohydrate molecules that project from other cell surfaces.

Question 26

When an antibody binds to its antigen, what are the three possible outcomes of this interaction?

Answer 26

1. Opsonization
2. Agglutination
3. Neutralization

• Opsonization is the marking of the pathogen for destruction by other white blood cells immediately
• Agglutination is the clumping together of the antigen and antibody into large insoluble protein complexes that can be phagocytized and digested by macrophages.
• Neutralization is making the pathogen or toxin unable to exert its effct on the body by neutralizing the antigen (preventing the antigen from binding to its target)

Question 27

What are the relative benefits of native-PAGE compared to SDS-PAGE?

Answer 27

Native-PAGE allows a complete, functional protein to be recovered after analysis, and more accurately determines the relative globular size of proteins.

• Only true if stain is not used, because stains typically denature proteins.

SDS-PAGE can be used to eliminate conflation from mass to charge ratios by analyzing only the mass of a protein.

• Sodium dodecyl phosphate (SDS) is a detergent that denatures the protein by disrupting noncovalent interactions and gives the protein-SDS complex a uniformly negative charge.

Question 28

What are the two potential drawbacks of affinity chromatography?

Answer 28

The two potential drawbacks of affinity chromatography are:

1. The protein of interest may not elute from the column because its affinity to its receptor/target is too high
2. The protein of interest may be permanently bound to the free receptor in the eluent.

Question 29

Why are proteins analyzed after isolation?

Answer 29

Protein isolation is generally on the first step in an analysis. The protein identity must be confirmed by amino acid analysis or activity. With unknown proteins, classification of their features is generally desired.

Question 30

What factors would cause a protein activity assay to display lower activity than expected after protein concentration is determined?

Answer 30

Contamination of a sample with a detergent or SDS could yield an artificially increased protein level, leading to lower protein activity than expected.

Alternatively, the enzyme could have denatured during isolation and/or analysis.