Protein: Structure + Function (lec 14 not on midterm 2)

Question 1

(T/F) Human genome has around 20k to 23k protein encoding genes that through alternative splicing of mRNAs and post-translational modifications generate hundreds of thousands of distinct protein activity.

Answer 1

True!

Question 2

What are proteins?

Answer 2

Polymers composed of the 20 amino acids with different R group side chains.

Amino acid side chains determine the distinct properties of individual proteins.

Question 3

Match the following protein structures to their definition:

1) Primary
2) Secondary
3) Tertiary
4) Quaternary
5) Supramolecular complexes

A) large-scale assembly - consisting of tens to hundreds of subunits and other biopolymers such as nucleic acids (big power house engines; RIBOSOME)

B) association between multipeptide complexes (multimeric structure)

C) local folding of the chain into alpha helices and b sheets with lots of H-bonds.

D) overall 3D conformation - composed of 2° structural elements and loops + turns. May form distinct, independently stable domain.

E) linear sequence of amino acids linked by peptide bonds

Answer 3

Primary: linear sequence of amino acids linked by peptide bonds

Secondary: local folding of the chain into alpha helices and b sheets with lots of H-bonds.

Tertiary: overall 3D conformation - composed of 2° structural elements and loops + turns. May form distinct, independently stable domain.

Quaternary: association between multipeptide complexes (multimeric structure)

Supramolecular: large-scale assembly - consisting of tens to hundreds of subunits and other biopolymers such as nucleic acids (big power house engines; RIBOSOME)

Question 4

What does protein function depend on?

Answer 4

Specific binding interactions and conformation changes in the structure of a properly folded protein

Question 5

What are the 6 main functions of proteins? Briefly describe them.

Answer 5

1) Structure - they organize the genome, organelles, cytoplasm, protein complexes, and membranes in 3D space

2) Regulation - control protein activity (e.g. post translation modification)

3) Signalling - monitoring the environment and transmitting information

4) Transport - moving small molecules and ions across membranes

5) Enzyme activity - catalyzing chemical reactions

6) Motors - generating force for movement (flagella, actin filaments, etc)

Question 6

How is a peptide bond formed?

Answer 6

Formed by a DEHYDRATION reaction linking one amino acid C-terminus to another amino acid N-terminus.

Question 7

(T/F) In a linear polymer (polypeptide), there is a free amino end (N-terminus) and a free carboxyl end (C-terminus).

Answer 7

True!

Question 8

Fill in the blank:

Protein function is derived from the ____ structure, which is determined by the aa sequence and ________ reactions.

Answer 8

3d; intramolecular

Question 9

Define secondary structures.

Answer 9

Stable spatial arrangements of polypeptide chain segments held together by H bonds between backbone amine and carbonyl groups.

Alpha helix and beta sheets.

Question 10

Which one of the statements regarding alpha helix is false?

1) They occur when the polypeptide backbone (ribbon) folds into a spiral/helix

2) Helix is stabilized by hydrogen bonds between backbone oxygen and hydrogen atoms (more bonds, more stable)

3) R groups project outward from the surface of the helix and determine the chemical nature of helix faces

4) Prolines are a major component of the alpha helix

Answer 10

4! Prolines cannot participate in H bonding and are usually excluded from alpha helix because they form kinks. Kinks constraints alpha helix from spiral.

Question 11

What is a beta sheet? How is it stabilized?

Answer 11

LATERALLY packed beta strands, each of which is a nearly fully extended polypeptide segment.

Stabilized by H bonds between backbone oxygen and hydrogen atoms in amino acids on DIFFERENT STRANDS.

Question 12

What is the difference between Antiparallel and Parallel beta sheets?

Answer 12

In parallel beta-sheets the strands all run in one direction (N-to-C), whereas in antiparallel sheets they all run in opposite directions.

Question 13

What produces a pleated polypeptide backbone contour?

Answer 13

Alpha carbon bond angles

Question 14

(T/F) In β sheets, alternate R groups project above and below the plane of the sheet.

Answer 14

True!

Question 15

Fill in the blanks regarding a β turn.

A β turn is composed of _____ residues.

It ______ the direction of a polypeptide chain ( ____ U-turn)

Cα carbons of the first and the fourth residues are usually less than ____ nm apart and linked by a ______ bond.

Answer 15

four

reverses; 180°

0.7; hydrogen

Question 16

What does the β turn facilitate?

Answer 16

β turns facilitate the folding of long polypeptides into compact structures.

Question 17

Which amino acids are commonly found in β turns?

Answer 17

Glycine (smallest R group) and Proline (built in bend)

Question 18

What are structural motifs? What can they be encoded by?

Answer 18

Structural motifs are regular combinations of secondary structures usually with a specific type of function.

They can be encoded by highly conserved sequence motif.

Question 19

What are the three types of structural motifs?

Answer 19

1. Coiled-coil motif
2. EF hand/helix-loop-helix motif
3. Zinc-finger motif

Question 20

Answer the following questions regarding coiled-coil motif:

1) Briefly describe the structure.

2) At which positions do the alpha helix-heptad repeat sequence have a hydrophobic residue?

3) How is the seam stabilized?

Answer 20

1. Two alpha helices that wound around each other. The hydrophilic molecules are outside, hydrophobic molecules are inside.
2. At positions 1 and 4, there is a hydrophobic residue.
3. Seam is stabilized by interactions between heptad 1 and 4 hydrophobic side chains!

Question 21

Where is zinc-finger motif present?

Answer 21

Zinc-finger motifs are present in many DNA-BINDING PROTEINS that help regulate TRANSCRIPTION.

Question 22

Fill in the blanks regarding zinc-finger motif:

It is composed of ____ amino acids with two INVARIANT _______ residues at position 3 and 6 and two INVARIANT _______ residues at position 20 and 24.

Answer 22

25; cysteine; histidine

*invariant means highly conserved; if you lose one of them, the motif can’t be formed.

*all organisms have these cys and his exactly in the same position.

Question 23

Cysteine and histidine interact to bind to Zinc(2+) between:

Answer 23

a pair of beta strands and a single alpha helix

Question 24

How is the tertiary structure stabilized?

Answer 24

1) Hydrophobic interactions between non-polar side chains.

2) Hydrogen bonds involving polar side chains and backbone amino and carboxyl groups.

Question 25

What happens to the hydrophobic residues in the tertiary structure?

Answer 25

They cluster together like drops of oil in the folded protein core, driven away from the aqueous surroundings by the HYDROPHOBIC EFFECT.

Question 26

What happens to the hydrophilic residues in the tertiary structure?

Answer 26

They (charged + uncharged polar side chains) form stabilizing interactions with surrounding water and ion on the protein surface.

Question 27

Fill in the blanks:

In the hemagglutinin (HA) tertiary structure, the distal domain (HA2) is the _______ domain and the proximal domain (HA1) is the _______ domain.

Answer 27

Globular; fibrous

Question 28

What does the hemagglutinin (HA) quaternary structure compose of?

How is it stabilized?

Answer 28

Three HA1-HA2 subunits.

It is stabilized by LATERAL INTERACTIONS between the long helices, forming a triple-stranded COILED-COIL stalk.

Question 29

What and where do the distal globular domain of the hemagglutinin quaternary structure bind to?

*hemagglutinin is an influenza virus surface protein

Answer 29

Sialic acid on the surface of target cells.

Question 30

(T/F) Approximately 20 percent of eukaryotic proteins have multiple structural domains.

Answer 30

False! Approximately 75 percent of eukaryotic proteins have multiple structural domains.

Question 31

Match the following terms to their definition regarding epidermal growth factor (EGF) domains:

1) EGF precursor
2) Neu
3) Tissue plasminogen activator (TPA)

A) Composed of EGF and other domains and breaks down blood clots

B) Generated by proteolytic cleavage that generates multiple EGFs

C) EGF domain plus another domain and is important for neural development

Answer 31

EGF precursor: Generated by proteolytic cleavage that generates multiple EGFs

Neu: EGF domain plus another domain and is important for neural development

Tissue plasminogen activator (TPA): Composed of EGF and other domains and breaks down blood clots

Question 32

(T/F) A protein can compose of multiple domains, while some domains are for structural activity other domains are for metabolic activity.

Answer 32

True!

Question 33

What is an example of a supramolecular complex?

Answer 33

Transcription initiation complex!

It is a molecular machine: Core RNA polymerase + general transcription factors, a mediator complex containing about 20 subunits and other protein complexes assemble at a gene promoter.

Question 34

Fill in the blank:

Protein function depends on binding ______.

Answer 34

Ligands (other molecules)

Question 35

How do enzymes accelerate rates of cellular reactions?

Answer 35

1. Lower activation energy
2. Stabilize transition state intermediates

Question 36

(T/F) Metabolic pathway enzymes may be associated as domains of a monomeric protein, subunits of a multimeric protein, or components of a protein complex assembled on a common scaffold.

Answer 36

True!

Question 37

(T/F) In all enzymes, like Trypsin, the catalytic site and the substrate-binding pocket are structurally distinct.

Answer 37

False!

In some enzymes, the catalytic site and the substrate-binding pocket OVERLAP.

Question 38

What side chains does the catalytic site of the trypsin contain?

Answer 38

Contains the side chains of the CATALYTIC TRIAD; Ser-195, Asp-102, and His-57, that BREAKS peptide bonds.

*some enzymes break peptide bonds, while others make.

Question 39

What are the functions of serine proteases?

Answer 39

1) Digest meals (tyrpsin, chymotyrpsin + elastase)

2) Control blood clotting (thrombin)

3) Help silk moths chew their way out of their cocoons (cocoonase)

Question 40

Fill in the blank:

Enzyme’s substrate binding site and catalytic site cooperate in _________ reaction to convert substrate to product.

Answer 40

Multistep

Question 41

What is the enzyme-substrate complex in equilibrium with?

Answer 41

EQUILIBRIUM with the unbound enzyme and substrate.

Question 42

The enzyme-substrate complex is the __________ step in the conversion of substrate to product.

Answer 42

INTERMEDIATE

Question 43

What does protein degradation do?

Answer 43

1. Regulates the life spans of intracellular proteins (short as a few mins for mitotic cyclins to as long as the age of an organisms for proteins in the lens of the eye).
2. Removes damaged proteins.

Question 44

Match the following steps of POLYUBIQUITINYLATION targeting proteins for PROTEASOMAL DEGRADATION:

1) Step 1
2) Step 2
3) Step 3
4) Step 4

A) E1 transfers the Ub molecule to a cysteine residue in E2.

B) E3 adds Ub molecules to the previously added Ub on the target protein

C) Enzyme E1 is activated by attachment of a ubiquitin (Ub) molecule

D) Ubiquitin ligase (E3) transfers the E2-bound Ub molecule to the side-chain - Nh2 of a lysine residue in a target protein, forming an isopeptide bond

Answer 44

Step 1: Enzyme E1 is activated by attachment of a ubiquitin (Ub) molecule

Step 2: E1 transfers the Ub molecule to a cysteine residue in E2.

Step 3: Ubiquitin ligase (E3) transfers the E2-bound Ub molecule to the side-chain - Nh2 of a lysine residue in a target protein, forming an isopeptide bond

Step 4: E3 adds Ub molecules to the previously added Ub on the target protein

Question 45

Match the final steps of ubiquitin and proteasome mediate proteolysis:

1) Recognition
2) Release
3) Unfolding
4) Transfer
5) Cleavage
6) Discharge

A) Deubiquitinase enzyme removes the Ub groups

B) A pore in the hexamer transfers the unfolded protein into the core proteolysis chamber.

C) Proteasome 19s cap Ub receptors bind the polyubiquitinylated target.

D) Cleaved peptides (2-24 residues long) are discharged and degraded by soluble proteases.

E) ATP hydrolysis enables the six-protein (hexameric) ATPase subunits to unfold the substrate.

F) Peptide bonds are cleaved.

Answer 45

1) Recognition: Proteasome 19s cap Ub receptors bind the polyubiquitinylated target.

2) Release: deubiquitinase enzyme removes the Ub groups

3) Unfolding: ATP hydrolysis enables the six-protein (hexameric) ATPase subunits to unfold the substrate.

4) Transfer: A pore in the hexamer transfers the unfolded protein into the core proteolysis chamber.

5) Cleavage: Peptide bonds are cleaved.

6) Discharge: Cleaved peptides (2-24 residues long) are discharged and degraded by soluble proteases.

Question 46

What are kinases and phosphotases?

Answer 46

Kinases phosphorylate proteins. Phosphotase de-phosphorylate proteins.

Both actions can activate some proteins and inactivate other types of proteins.

Question 47

(T/F) Most kinases can phosphorylate multiple different target proteins, and human genome encodes around 500 kinases (kinome).

Answer 47

True!

Question 48

(T/F) Kinome can only phosphorylate certain proteins.

Answer 48

False! They can phosphorylate members of all protein classes.