Team:KAIST-Korea/Project/Modeling

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== Single chain antibody structural alignment ==
== Single chain antibody structural alignment ==
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[[Image:SingleChainAntibodyProtocol.png|thumb|500px|center|Single chain antibody structural alignment protocol]]
=== Protocol ===
=== Protocol ===
There are four steps to compare structure of single chain antibody and FGF binding domain of FGFR. First step is taking variable region sequences of antibodies. Next step is combining these variable region sequences with linker sequence to make single chain antibody sequence. Third step is predicting the structure of single chain antibody with structure prediction program like modeler. Final step is to structural align these structures of antibodies with structure of FGF binding domain of FGFR(PDB ID : 1EVT).
There are four steps to compare structure of single chain antibody and FGF binding domain of FGFR. First step is taking variable region sequences of antibodies. Next step is combining these variable region sequences with linker sequence to make single chain antibody sequence. Third step is predicting the structure of single chain antibody with structure prediction program like modeler. Final step is to structural align these structures of antibodies with structure of FGF binding domain of FGFR(PDB ID : 1EVT).

Revision as of 05:08, 11 August 2010

 

Modeling



Single chain antibody structural alignment

Single chain antibody structural alignment protocol

Protocol

There are four steps to compare structure of single chain antibody and FGF binding domain of FGFR. First step is taking variable region sequences of antibodies. Next step is combining these variable region sequences with linker sequence to make single chain antibody sequence. Third step is predicting the structure of single chain antibody with structure prediction program like modeler. Final step is to structural align these structures of antibodies with structure of FGF binding domain of FGFR(PDB ID : 1EVT).

Data source

Single chain antibody is the combination of variable regions of known antibodies with linker sequence which can bind to the antigens. We need to know the VL and VH sequences to make single chain antibody. The source of these antibody sequences are NCBI, Uniprot and RCSB PDB. NCBI and Uniprot provide the single chain sequence of variable regions (VL and VH) and antigen binding fragments (Fab). RCSB provide the structure of the complexes of antigenbinding fragment which binds to its antigens. But we only need the sequence of variable region. So we get the last 120~150 reside and assume them as the variable region. And data from RCSB contain not only sequence of antibody, but also antigens. S we filter them based on label of files.to get heavy chains and light chains of antibody.

Single chain antibody synthesis

We combine the antibody variable region sequences in order of VH-linker-VL to make single chain antibody sequence. The sequence of linker is GGGGSGGGGS

Structure Prediction

We used the Modeller program to predict the structure of single chain antibody from its sequence. Modeller predict 3D structure of protein with structure of know similar proteins based on homology model. Input file is the sequence of single chain antibody with fasta format and output file is the structure of single chain antibody with pdb format.

Structure Alignment

In this step, we check the structural similarity between single chain antibodies and FGF binding domain to align the structure of single chain antibody with FGF binding domain of FGFR. The structure of FGF binding domain of FGFR is provided by RCSB PDB(PDB ID : 1EVT) We used Matt structural alignment program to do this job. Matt do the structural alignment which minimize the distance between α-carbon chain of two proteins based on the common structure (α helix). Input file is the structure of single chain antibody with pdb format and output file is the text file which contain the number of amino acids which compose shared structure (Core residue), average distance between alpha carbon chains of two proteins (Core RMSD), the score of similarity which is calculated by Matt(Raw score) and the probability that this similarity is just the product of random(p-value) and pdb files which contain the alignment result of single chain antibody with FGF binding domain of FGFR.

Result : Table

Name Core ResiduesCore RMSDRaw ScoreP-value
2VXT 95 2.676 84.306 0.000222
2VXU 98 2.563 92.118 0.0000823
2VXV 77 2.64 52.269 0.0644
2ZKH 96 2.855 93.411 0.001
3AAZ 84 4.197 68.078 0.0642
3D69 137 12.224 98.134 0.00541
3EO9 80 2.282 84.665 0.00295
3EOA 116 10.402 98.544 0.0547
3EOB 68 4.685 56.357 0.3753
3EYV 85 4.704 66.669 0.1653
3FMG 81 2.629 73.298 0.0129
3FOG 90 7.243 74.031 0.0785
3G6D 72 4.403 69.774 0.1281
3GBM 119 10.331 115.929 0.0314
3GBN 77 2.788 57.204 0.0592
3GHB 101 12.861 81.189 0.3984
3GHE 96 3.302 109.208 0.00117
3GI8 127 6.837 113.467 0.00038
3GI9 52 3.534 43.56 0.166
3GIZ 89 8.217 68.702 0.3172
3GK8 91 6.84 84.048 0.00811
3GKW 74 3.205 47.096 0.1215
3GNM 135 4.739 103.812 0.00000297
3GO1 103 8.842 77.631 0.2587
3GRW 107 9.819 108.405 0.0449
3H42 69 4.389 59.165 0.3421
3HC0 67 3.065 40.982 0.1028
3HC3 67 3.173 49.109 0.0944
3HC4 36 1.998 42.538 0.206
3HI5 77 3.222 58.096 0.1832
3HI6 81 3.646 63.526 0.0394
3HMW 83 4.159 62.598 0.1976
3HMX 70 5.215 62.814 0.2228
3HNT 84 2.62 74.666 0.00118
3HNV 84 2.998 77.17 0.00374
3HR5 78 4.158 58.026 0.2978
3I50 65 2.206 52.252 0.1156
3I9G 84 5.026 64.155 0.1506
3IU3 75 3.192 56.006 0.2552
3IXT 78 2.951 68.627 0.0378
3KDM 87 4.273 65.722 0.1683
3KS0 81 3.039 55.562 0.0192
3KYK 88 4.728 68.681 0.1016
3KYM 85 6.52 66.102 0.5135
3L1O 93 2.545 85.909 0.0014
3L5W 89 4.925 72.178 0.0637
3L5X 88 4.796 65.262 0.1111
3L5Y 64 4.689 65.227 0.2007
3L95 73 5.105 70.312 0.0515
3LMJ 76 2.699 56.627 0.0237
3LQA 75 8.371 58.374 0.8315
3LS4 79 2.458 72.68 0.00694
3LS5 81 2.443 73.043 0.0014
3LZF 96 2.468 93.292 0.00155
3MLR 71 2.574 64.979 0.0165
3MLS 75 2.534 57.386 0.0202
3MLU 127 10.025 73.766 0.0305
3MLV 75 2.454 62.321 0.0218
3MLW 83 8.94 59.029 0.4846
3MLX 118 11.485 105.391 0.0961
3MLY 84 4.704 63.962 0.1023
3MLZ 82 3.995 64.563 0.123
3MUG 79 2.827 68.72 0.0223
3MXV 125 10.857 121.911 0.01
3MXW 102 11.262 78.348 0.4213
16A1 86 2.139 89.893 0.00443
  • Name -- Except 16A1(Name of antibody itself), all name of single chain antibodies are came from PDB ID of its source.;



Result : Figures

Using arranged pdb files and protein 3D structure drawing program - PyMOL, again, we predict structural similarlity between the structure of FGF binding domain and single chain antibodies associated with the table. In order to show the structures clearly, we control the shape setting 'cartoon' and 'chain'. Cyan Color is the structure of FGF binding domain and Green Color is the comparative antibodies. In the result, we can easily see all antibodies' structure are very similar completely or symmetrically.



Result Analysis

The table above shows the result of alignment of 65 single chain antibody for control group and 1 single chain antibody for our real experiment (16A1). Higher core residue and Raw score and lower core RMSD and p-value means more similar. 16A1 antibody shows 26th highest in core residue, 2nd lowest in core RMSD, 13th highest Raw score and 13th lowest p-value. These result means that our 16A1 antibody has higher similarity than average antibodies.