Modular proteins

Question 1

What is a domain?

Answer 1

a folded structural unit that need not be contiguous. e.g. MHCII

Question 2

what is a. module?

Answer 2

a domain with a contiguous sequence. repeately used in diverse protiens. e.g. F3

Question 3

what is a repeat?

Answer 3

a unit htat does not fold in isolation. several copies are needed. e.g. Leu-rich repeat/

Question 4

what are some of the biological implications of protein domains?

Answer 4

Many surfaces use the same scaffold.
presentation of binding sites. can be in many palces

Question 5

describe the structure of EGF and its amino acid information?

Answer 5

has about 40 residues and 3 disulphide bonds. it is a two stranded beta sheet followed by a loop to a C terminal short two stranded sheet.

its subdomains between the conserved cystines vary in length.

Question 6

describe the structure of calcium binding EGF (cbEGF) and its amino acid information?

Answer 6

3 disulphide bonds. two stranded beta sheet followed by a loop to a C terminal short two stranded sheet.

is often as a tandem repeat and is the main building block for fibrillin.

Question 7

what is the domain organsiation of Fibrillin?

Answer 7

around 37 cbEGF
Tb dominan
a few non cb EGF

some hybrid domains:
unique region at N terminus
a proline rich region

Question 8

what is the function of fibrillin?

Answer 8

forms microfibrils that are part of the ECM. 8 strands of fibrilin come together to form a microfibril.

Question 9

mutations of fibrillin are associated with what disease?

Answer 9

more than 500 mutations on fibrillin and are all assocaited with a class of diseases called Marfan syndrome.

Question 10

What are symptoms of mild and severe marfan syndrome?

Answer 10

tall and lanky when mild

severe marfans shows CV problems, vision problems, and leathal at developmental stage.

Question 11

What are the two models for microfibril strucuture?

Answer 11

Jack knife model and antiparralel staggered model.

Question 12

What is the jack knife model?

Answer 12

Fibrillin aranged in a jack knmife conformation and then these are arranged ontop of each other – there is growing evidence that this is the correct model

Question 13

what is the antiparralel staggared conformation model?

Answer 13

fibnrillins layed out partially overlapping generating a repetative pattern with a 50nm register

Question 14

What joins cbEGF- cbEGF junctions?

Answer 14

a 1 residue linker that has a well coserved interface. they are well ordered

Question 15

what is found at a cbEGF-Tb juntion?

Answer 15

a variable linker that has a not conserved interfance. are shown to be flexible.

Question 16

what is multivalency?

Answer 16

the ability of a ligand to bind to multiple binding sites on a protein or receptor simultaneously.

Question 17

What is a monovalent ligand?

Answer 17

a ligand that has only one binding site.

Question 18

What is a bivalent ligand?

Answer 18

composed of two functional pharmacophores linked by a spacer. are expected to bind to and stabilise preexisiting dimers.

Question 19

Antibodies are examples of polyvalent interactions. what are examples of Igs that are bivalent, tetravalent and decavalent?

Answer 19

Bivalent: IgD, IgE, IgD
Tetravalent: IgA
Decavalent: IgM

Question 20

What is fibronectin

Answer 20

a type of glycoprotein that plays importnatn roles in various biological processes.

Question 21

what are the two forms of fibronectin

Answer 21

it exists in two forms: circulating in the blood and activated in the extracellular matrix (ECM)

Question 22

what is the size of fibronectin

Answer 22

approx 250 kda and 70 nm in size

Question 23

what are the functions of fibronectin?

Answer 23

involved in cell adhesion, cell migration, wound healing, cancer progression and development.

Question 24

what happens in the absence of fibronectin

Answer 24

gene knockout studies have shown that fibronectin is essential for embryonic development and its absence is fatal.

Question 25

what are the specific binding regions in fibronectin called?

Answer 25

Motifs

Question 26

How does the binding affinity of fibronectin change when the polycalency of the binding region is increased?

Answer 26

The binding affinity of fibronectin increases when the polyvalency of the binding region is increased.

Question 27

How long are the typical binding motifs in fibronectin?

Answer 27

They are around 8 ammino acids long.

Question 28

Are the binding motifs in fibronectin conserved?

Answer 28

they are almost conserved in multiple places

Question 29

How many fibronectins can bind to one molecule by aligning the run of 4 amino acids multiple times?

Answer 29

by allgiing the run of 4 amino acids multiple times, up to 11 fibronectins can bind to one molecule.

Question 30

What is the role of fibronectin in bacterial uptake?

Answer 30

Fibronectin plays a role in mediating bacterial uptake by epithelial or endothelial cells.

Question 31

What are the fibronectin binding repeats (FnBR) and how do they bind to fibronectin?

Answer 31

The fibronectin binding repeats (FnBR) are sequences that bind to fibronectin via 4 to 5 N-terminal F1 domains. Multiple fibronectin molecules can bind depending on the number of FnBR in the bacterial receptor, which leads to the formation of a second layer of polyvalency.

Question 32

How does clustering of RGD sequences on fibronectin activate integrins?

Answer 32

Clustering of RGD sequences on fibronectin activates integrins by binding to their ligand-binding domain.

Question 33

What is the role of integrins in fibronectin-mediated bacterial uptake?

Answer 33

Integrins play a crucial role in fibronectin-mediated bacterial uptake by clustering on the host cell and allowing the bacteria to be engulfed.

Question 34

Why is it important for the bacteria to hide from the immune system?

Answer 34

It is important for the bacteria to hide from the immune system because persistent infection can occur unless the bacteria has somewhere to hide. Thus, it hijacks the migration capability of the host cell to be engulfed and hide from the immune system.

Question 35

What is the SH2 domain?

Answer 35

The Src homology 2 (SH2) domain is a protein domain of approximately 100 amino acids that recognizes and binds to specific phosphorylated tyrosine residues on target proteins.

Question 36

What is the structure of the SH2 domain?

Answer 36

The SH2 domain has a central five-stranded beta-sheet and two flanking alpha-helices, forming a “socket” with two holes to plug in a peptide. The domain’s backbone is an anti-parallel sheet formed by strands A, B, C, D, and G.

Question 37

What is the primary function of the SH2 domain?

Answer 37

The primary function of the SH2 domain is to recognize and bind to specific phosphorylated tyrosine residues on target proteins.

Question 38

How does the SH2 domain recognize target sequences?

Answer 38

The SH2 domain recognizes target sequences that have a two-pronged plug configuration, with a phosphorylated tyrosine followed by specific amino acids.

Question 39

What are the two functionally distinct sides of the SH2 domain?

Answer 39

One side of the SH2 domain, flanked by helix A, is primarily involved in binding to phosphotyrosine residues, while the other side, flanked by helix B and the EF and BG loops, interacts with side-chains of the peptide that are C-terminal to the phosphotyrosine.

Question 40

How many SH2 domains are there in the human complement?

Answer 40

The human complement of SH2 domains comprises 120 domains found in 110 different proteins, including signaling proteins like Src, BLNK, and CNI.

Question 41

Why are SH2 domains important?

Answer 41

The SH2 domain plays a crucial role in regulating protein-protein interactions and signaling pathways, making it an important target for drug discovery and development.

Question 42

What is the SH3 domain?

Answer 42

The Src homology 3 (SH3) domain is a protein domain of approximately 60-70 amino acids that recognizes and binds to specific proline-rich motifs on target proteins.

Question 43

What is the structure of the SH3 domain?

Answer 43

The SH3 domain has a fold consisting of two perpendicular anti-parallel beta-sheets flanked by two alpha-helices, forming a “barrel” shape.

Question 44

How does the SH3 domain recognize target sequences?

Answer 44

The SH3 domain recognizes target sequences that have a proline-rich motif, with an aromatic or acidic residue at the P-3 position. The domain has a wide and shallow binding groove that accommodates the proline-rich ligand. (PXXP motif)

Question 45

what is the domain arrangement of an inactive Src Kinase?

Answer 45

Kinase domain - with N lobe, C lobe and ATP (analogue) wedged in-between

SH3 domain, SH2 domain wedged on back of kinase - this is inactive state - very low activity due to this

Purpose of the kinase is to phosphorylate tyrosine by using the tertiary phosphate from ATP

Question 46

what keeps Src kinases inactive>

Answer 46

SH3 domain interacts with a poly proline motif pXXp on the linker between SH3, SH2 and the kinase domain

Also in the case of Src - C-terminal tail has a phosphorylated tyrosine that plugs into the SH2 domain

Question 47

How are Src kinases activated?

Answer 47

by a process called switching.

a polyproline binds to the SH3 and SH2 of the Src kinase and causes them to unlatch. unlatching is helped by depophorylation of tyr 527.

allows the N and C lobes of the kinase to move apart and become free to function. enhances the catalytic rate, in doing so the ATP will use a tertiary phosphate to phosphorylate the tyrosine on the activation loop (Tyr416)

Once this is phosphorylated and activation loop is fully activated - it is switched on (this is the switching process)