Block B lecture 3

Question 1

what do antigens bind to a complex of for T cells ?

Answer 1

T-cell receptors (TCRs) bind a complex of an antigen fragment and a self molecule ( MHC)

Question 2

how are T cell receptors different to antibodies on recognising epitopes ?

Answer 2

Unlike most antibodies, TCRs can recognise epitopes that are buried within antigens These antigens must first be degraded by proteases into fragments and the peptide epitope delivered to a self molecule, called an MHC (major histocompatibility complex) molecule . It is in this form, as a complex of peptide and MHC molecule, that antigens are recognized by TCRs .

Question 3

what does the TCR resemble ?

Answer 3

The TCR resembles a membrane-bound Fab fragment ( antibody). The Fab fragment of an antibody molecule is a disulphide-linked heterodimer, each chain of which contains one antibody C domain and one V domain; the juxtaposition of the V domains forms the antigen-binding site.

The TCR is also a disulphide-linked heterodimer, with each chain containing an antibody C- like domain and an antibody V-like domain. As in the Fab fragment, the juxtaposition of the V domains forms the site for antigen recognition.

Question 4

describe the structure of the TCR?

Answer 4

The TCR heterodimer is composed of two transmembrane glycoprotein chains, α and β. The extracellular portion of each chain consists of two domains, resembling antibody V and C domains, respectively. Both chains have carbohydrate side chains attached to each domain.

A short stalk segment, analogous to an antibody hinge region, connects the Ig-like domains to the membrane and contains the cysteine residue that forms the interchain disulfide bond. The transmembrane helices of both chains are unusual in containing positively charged (basic) residues within the hydrophobic transmembrane segment. The α chain carries two such residues; the β chain has one.

Question 5

how do B cells interact with an antigen ?

Answer 5

B cells bind directly to intact antigens, typically interacting with the surface of proteins, contacting amino acids that are discontinuous in the primary structure but are brought together in the folded protein.

Question 6

T cells interact with antigen ?

Answer 6

T cells interact with short continuous amino acid sequences, which may be buried within the protein.

• Thus, proteins must be unfolded and digested into fragments to be recognised by T cells
• T cells recognise such fragments only when presented by MHC molecules

Question 7

do T cells recognise antigenic proteins or peptide fragments ?

Answer 7

TCR recognises not the antigenic protein itself but a peptide fragment of the protein

Question 8

in the MHC class I molecules , does the alpha and beta chains both span the membrane ?

Answer 8

MHC class I molecule is a heterodimer of a membrane-spanning α chain bound non-covalently to β2- micro-globulin , which does not span the membrane

Question 9

what does the alpha 3 and beta 2 domain share similarities with ?

Answer 9

the α3 domain and β2-micro-globulin show similarities in amino acid sequence to antibody C domains and have similar folded structures.

Question 10

what about the alpha one and 2 domains ?

Answer 10

the α1 and α2 domains are part of the same polypeptide and fold together into a single structure consisting of two separated α helices lying on a sheet of eight antiparallel β strands. The folding of the α1 and α2 domains creates a long cleft or groove, which is the site at which peptide antigens bind to the MHC molecules. For class I molecules, this groove is open at only one end.

Question 11

MHC class II structure ?

Answer 11

MHC class II molecules resemble MHC class I molecules in overall structure. The MHC class II molecule is composed of two transmembrane glycoprotein chains, α and β . Each chain has two domains, and the two chains together form a compact four-domain structure similar to that of the MHC class I molecule

Question 12

what are the alpha 2 and beta 2 domains show similarities to MHC class II?

Answer 12

Alpha 2 and beta 2 domains of the MHC class II molecule, have amino acid sequence and structural similarities to antibody C domain

Question 13

How do MHC molecules bind peptides within the cleft ?

Answer 13

tightly

Question 14

how is the peptide bound in MHC class I ?

Answer 14

In MHC class I molecules the peptide is bound in an elongated conformation with both ends tightly bound at either end of the cleft

Question 15

how is the peptide bound in the MHC class II molecule ?

Answer 15

MHC class II molecules , the peptide is also bound in an elongated conformation but the ends of the peptide are not tightly bound and the peptide extends beyond the cleft

Question 16

how is the upper surface of the MHC peptide complex recognised by ?

Answer 16

The upper surface of the peptide:MHC complex is recognized by T cells and is composed of residues of the MHC molecule and the peptide

Question 17

what happens to the amino acid side chains of the peptide ?

Answer 17

the amino acid side chains of the peptide insert into pockets in the peptide-binding groove of the MHC molecule; these pockets are lined with residues that are polymorphic within the MHC.

Question 18

how are the polypeptides bound to the MHC class I molecule ?

Answer 18

Polypeptides are bound to MHC class I molecules by their ends

Question 19

how does the MHC class I interact with the backbone of the bound peptide ?

Answer 19

MHC class I molecules interact with the backbone of a bound peptide through a series of hydrogen bonds and ionic interactions at each end of the peptide

Question 20

where does the amino terminus lie ?

Answer 20

The amino terminus of the peptide is to the left,

Question 21

carboxy terminus ?

Answer 21

the carboxy terminus to the right

Question 22

what does the cluster of tyrosine residues found in all MHC class I molecules ?

Answer 22

A cluster of tyrosine residues common to all MHC class I molecules forms hydrogen bonds to the amino terminus of the bound peptide

Question 23

the second cluster of residues form ?

Answer 23

while a second cluster of residues forms hydrogen bonds and ionic interactions with the peptide backbone at the carboxy terminus and with the carboxy terminus itself.

Question 24

how does the peptide bind to the MHC molecule 1 ?

Answer 24

peptides bind to MHC I molecules through structurally related anchor molecules. Peptides eluted from two different MHC class I molecules are shown in the upper and lower panels, respectively. The anchor residues (green) differ for peptides that bind different allelic variants of MHC class I molecules but are similar for all peptides that bind to the same MHC molecule. The anchor residues that bind a particular MHC molecule need not be identical, but are always related: for example, phenylalanine (F) and tyrosine (Y) are both aromatic amino acids, whereas valine (V), leucine (L), and isoleucine (I) are all large hydrophobic amino acids. Peptides also bind to MHC class I molecules through their amino (blue) and carboxy (red) termini

Question 25

peptides bind to MHC class II molecule ?

Answer 25

Peptides bind to MHC class II molecules by interactions along the length of the binding grove. A peptide (yellow; shown as the peptide backbone only, with the amino terminus to the left and the carboxy terminus to the right) is bound by an MHC class II molecule through a series of hydrogen bonds (dotted blue lines) that are distributed along the length of the peptide. The hydrogen bonds toward the amino terminus of the peptide are made with the backbone of the MHC class II polypeptide chain, whereas throughout the peptide’s length bonds are made with residues that are highly conserved in MHC class II molecules. The side chains of these residues are shown in grey on the ribbon diagram of the MHC class II groove.

Question 26

length of MHC class I and II peptide differ in length ?

Answer 26

MHC class II peptides tend to be longer in length , while MHC class I peptides are shorter.

Question 27

are the peptides that bind to MHC class II all the same length and have anchor regions at the same places ?

Answer 27

Peptides that bind MHC class II molecules are variable in length and their anchor residues lie at various distances from the ends of the peptide

Question 28

Hydrophobic region ?

Answer 28

Note that all of the peptides share a hydrophobic residue in position 1 and 9 [for example, tyrosine (Y), leucine (L), phenylalanine (F)]

Question 29

negative charge ?

Answer 29

, a negatively charged residue in position 4 aspartic acid (D) or glutamic acid (E)

Question 30

basic ?

Answer 30

[lysine (K), arginine (R), histidine (H), glutamine (Q), or asparagine (N)] in position 6

Question 31

explain how the TCR binds to the peptide and MHC complex ?

Answer 31

the TCR binds to the top of a peptide:MHC class I molecule, touching both the α1 and α2 domain helices. The CDRs of the TCR are shown in colour: the CDR1 and CDR2 loops of the β chain are light and dark blue, respectively; and the CDR1 and CDR2 loops of the α chain are light and dark purple, respectively. The α-chain CDR3 loop is yellow, and the β-chain CDR3 loop is green. The β-chain HV4 loop is in red (TCR chain contain a fourth hypervariable region located out with the antigen receptor). The thick yellow line P1–P8 is the bound peptide. Panel b: the outline of the TCR’s antigen-binding site (thick black line) is superimposed over the top surface of the peptide:MHC complex (the peptide is shaded dull yellow). The TCR lies at a somewhat diagonal angle across the peptide:MHC complex, with the α and β CDR3 loops of the T-cell receptor (3α, 3β, yellow and green, respectively) contacting the center of the peptide. The α-chain CDR1 and CDR2 loops (1α, 2α, light and dark purple, respectively) contact the MHC helices at the amino terminus of the bound peptide, whereas the β-chain CDR1 and CDR2 loops (1β, 2β, light and dark blue, respectively) make contact with the helices at the carboxy terminus of the bound peptide.

Question 32

how are the TCR alpha and beta chain genes composed of and how is diversity occurred?

Answer 32

The TCRα- and β-chain genes are composed of discrete segments that are joined by somatic recombination during development of the T cell in the thymus. Functional α- and β-chain genes are generated in the same way that complete immunoglobulin genes are created and with the aid of RAG-1/2.

Question 33

for the V alpha gene segment how does it rearrange?

Answer 33

a Vα gene segment rearranges to a Jα gene segment to create a functional V-region exon. Transcription and splicing of the VJα exon to Cα generates the mRNA that is translated to yield the TCR α- chain protein

Question 34

for the beta chain ?

Answer 34

For the β chain (lower part of figure), like the immunoglobulin heavy chain, the variable domain is encoded in three gene segments, Vβ, Dβ, and Jβ. Rearrangement of these gene segments generates a functional VDJβ V-region exon that is transcribed and spliced to join to Cβ; the resulting mRNA is translated to yield the TCR β chain

Question 35

when do the alpha and beta chains pair ?

Answer 35

The α and β chains pair soon after their synthesis to yield the α:β TCR heterodimer

Question 36

what is the role of RSS’s ?

Answer 36

RSSs flank TCR gene segments. As in the immunoglobulin gene loci, the individual gene segments at the TCRα and TCRβ loci are flanked by heptamer–spacer–nonamer RSSs and are recognised by the same enzymes. RSS motifs containing 12-bp spacers are depicted here as orange arrowheads, and those containing 23-bp spacers are shown in purple. Joining of gene segments almost always follows the 12/23 rule. Because of the disposition of heptamer and nonamer RSSs in the TCRβ and TCRδ loci, direct Vβ to Jβ joining is in principle allowed by the 12/23 rule (unlike in the antibody heavy-chain gene), although this occurs very rarely owing to other types of regulation that are still being investigated.

Question 37

does somatic hypermutation occur in T cells ?

Answer 37

Somatic hypermutation as a source of diversity is not included in this figure because it does not occur in T cells.

Question 38

where do TCR concentrate diversity.

Answer 38

TCRs concentrate diversity in the third hypervariable region and this interacts with the peptide of the peptide MHC complex.

Question 39

where does the CDR3 loops lie ?

Answer 39

CDR3 loops lie in the centre of the interface between the TCR and the peptide:MHC complex, and make direct contact with the antigenic peptide.