I. Introduction
II. General Structure
III. Loops on Light Chain
IV. Loops on Heavy Chain
V. Surface Topology
Myasthenia gravis (MG) is a condition which causes muscle weakness. In the neuromuscular junction, acetylcholine (ACh) is released and the ACh binds to receptors (AChR). MG is caused when the interaction between ACh and AChR fails because there is a loss of AchR. It has been shown that anti-AchR antibodies bind to the main immunogenic region (MIR) of AChR and cause their loss. The loss of AChR causes the symptoms seen in MG.
The Fab fragments were a part of the whole anti-AChR antibodies but have become detached. They contain part of the antigen binding region. As apposed to the whole anti-AChR antibodies, the Fab fragments do not cause loss of AChR. Instead Fab fragments are able to protect the AChR from loss due to anti-AChR antibodies. The Fab fragments bind to the MIR of the AChR and protect it from anti-AChR antibodies by not allowing them to bind. This protective quality is due to the univalence of Fab fragments. This means that Fab fragments bind in only one place, as apposed to the anti-AchR antibodies, which are bivalent. Fab192 binds to human AChR with unusual affinity. Through investigation of these Fab fragments, insight into possible treatments will be obtained.
The Fab192 fragment is made up of mostly beta sheets that can be see by pressing this button. < >. This image shows two copies of the Fab192 fragment one is magenta and the other is white. The Fab192 fragments are not bound together in nature as they are shown here. This head to tail conformation is merely the unit cell which is formed in crystallization. This model also includes 209 water molecules< >.
The Fab192 fragment shows the immunoglobulin fold which is the same for all members of the immunoglobulin super family of proteins. This region is composed of beta sheets and one disulfide bond. All immunoglobulins consist of a light chain and a heavy chain. For the Fab192 fragment the light chain< > and the heavy chain< > are shown. The light chain is the smaller of the two regions and is made up of 213 residues, while the heavy chain, the larger of the two regions contains 214 residues. Both chains have variable and nonvariable regions. The variable region is where foreign materials interact with the Fab fragment. The heavy and light chain each contain 3 loops that are responsible for interactions with AChR.
Reload PDB < >
The light chain has 3 loops that are involved with binding to the antigen, L1, L2, and L3. The loops are formed only in the hypervariable regions. <
L1 < > belongs to the canonical group 2. It contains the typical key residues L2 (Ile), L25 (Ala), L29 (Ile) and L71 (Phe)< >. L2 < > belongs to canonical group 1 and has key residues L48 (Ile) and L64(Glu)< >. L2 also contains a three residue hairpin turn made up of L50-L52 < > L3 < > does not fall into one of the canonical groups typically found in this region based on amino acid composition. It is only 5 amino acids long and is missing the proline usually found at either L94 or L95. However, in terms of its 3D structure the loop resembles canonical form 1. L90 (Gln) < > is the only key residue in fab 192. In the canonical form L93, L94, and L96 are also key residues.
Reload PDB < >
The heavy chain, like the light chain, also contains three loops, H1, H2, and H2, in the hypervariable region that are involved with binding to the antigen. < >. H1 < > falls into canonical group 1. It has key residues H26 (Gly), H27 (Phe), H29 (Leu), H34 (Val), and H97 (Arg) < >. Loop H2 < > belongs to canonical group 2 and is only three residues long. The three residues are part of a 7 residue turn. H2 has only one key residue H 55 (Gly) < >.
H3< > contains 10 residues and does not fit into a canonical group. H97(Arg) and H105(Asp) < > form an ion pair that has a large influence on the structure of the loop and causes a bulge. It has been shown that aromatics play an important role in antigen binding and about half of the residues on H3 are aromatic, therefore it can be assume that this region is significant for antigen binding .
This figure shows the electrostatic potential of the antigen binding site. Blue represents positively charged areas while red represents negatively charged areas. Fab192 is an antibody for a large protein antogen, therefore as would be expected the surface of the antigen-binding site is somewhat planar. The area of the binding site is 2865 Ǻ.
The region shown as a red strip down the center of the anitgen-binding site is where a cleft is formed. This cleft is bounded by side chains of residues H27 (Phe), H29 (Leu), H101 (Trp), and H104 (Phe). Residue H27 is found on loop H1, residue H101 is found on loop H3, and residue H104 is found on loop H3. The bottom of this cleft is negatively charged, as denoted by the red. This region is formed by the side chain residues H98 (Glu) and H99 (Asp) which are part of loop H3. This is where the cleft shown in red is located on the molecule < >
It can not be said for certain
where this region binds to the MIR region of AChR, but there are a number of
possible residues. There is an AChR sequence, which contains two acidic and
one basic residue. This region is known to participate in the epitopes of several
anti-MIR mAbs (67-76(WNPDDYGGVK)). In addition in the sequence of residues alpha77-87(KIHPSEKIWR)
the basic K76 is followed by four basic residues (K77, H79, K84, R87). There
is the possibility that these regions play critical roles in the epitope of
mAb192.
Kontou, Maria; Demetres D. Leonidas, Efstratia H. Vatzaki, Panayiota Tsantili, Avgi Mamalaki, Nikos G. Oikonomakos, K. Ravi Acharya and Socrates J. Tzartos. 2000. The crystal structure of the Fab fragment of a rat monoclonal antibody against the main immunogenic region of the human muscle acetylcholine receptor. European Journal of Biochemistry 267: 2389-2396.
http://www.tulane.edu/~biochem/med/igg.htm
http://stingray.bio.cmu.edu/~web/bc1/ig/ig.htm