Developed@Bioinformatics Centre, University of Pune, INDIA
The identification of the regions of
interaction between an antigen
(Ag) and an antibody (Ab) is one of the most interesting problems in
molecular immunology. Antigen-antibody (Ag-Ab) complexes are
non-obligatory heterocomplexes that are made and broken according to
the environment or the external factors and involve proteins (antigen
and antibody) that must also exist independently. The
most remarkable feature of antigen-antibody interactions is the high
affinity and strict specificity of antibodies for their antigens. It is
known that antibodies recognize the unique conformations and spatial
locations on the surface of antigens. Therefore,
are defined as the portions of the antigen molecules, which are
responsible for specificity of the antigens in Ag-Ab reactions and that
interact with the antigen binding site of antibody (paratope) to which
they are complementary.
Types of Epitopes
Both humoral and cellular arms of immune system
recognise and react
with only specific regions of the pathogen, called epitopes. It has
been known that B and T cells recognize different epitopes on the same
antigenic molecule. Thus, there are two types of
epitopes, B cell epitope and Th cell epitope. The B cell epitopes
are accessible, hydrophilic regions and majority of them are capable of
neutralisation. Antibodies produced by B cells recognise the
intact antigen in its native conformation. The epitopes
recognized by T cells are products of processed or partially degraded
proteins that are bound to MHC molecules and are usually amphipathic
(i.e., alternating hydrophobic and hydrophilic) regions.
The B cell
be contiguous / Sequential (when Ab binds to a contiguous stretch of amino acid residues
that are linked by peptide bond) or non-contiguous / Conformational (when Ab binds to non-contiguous residues, brought together
by folding of polypeptide chain). The specificity of sequential
epitopes (SE) is determined by the sequence of subunits (e.g. amino
acids). On the other hand, specificity of conformational epitopes (CE)
depends on the spatial folding or conformation of the contributing
individual sequential epitopes.
Existing Approaches to predict B-cell
& Woods (1981)
& Doolittle (1982)
et al. (1985)
et al. (1986)
& Tongaonkar (1990)
- The only
algorithm for the prediction of conformational epitopes is CE (Kolaskar & Urmila Kulkarni, 1999)
percent accessible surface area (%ASA) of each amino acid residue
is calculated using an implementation of Voronoi procedure (1908) by
McConky etal (2002) which is consistent with Lee
& Richards (1971).
having %ASA >= 25% are termed as accessible residues.
contiguous stretch of more than three accessible residues is termed as
antigenic determinant is extended to N- and C-terminals, only if,
accessible amino acid(s) were present after an inaccessible amino acid
residues which have %ASA>=25% and does not satisfy above criterion
are also taken into
antigenic determinants are then listed with details like the chain ID,
length of determinant, start and end position and the amino acid
amino acid residues that are part of antigenic determinants are shown
distance between every atom of residues from the ith determinant and
every atom of residues
from the jth determinants is calculated.
- If the
distance between any pair of atoms of at
least two residues from sequentially distinct determinants was found to
be <= predetermined
cutoff distance, then jth determinant was termed to be part of a
conformational epitope that
consists of ith and jth determinants.
residues which are not part of AD and satisfy
distance and %ASA criteria are also included as a part of CE.
were carried out for every sequential determinant (j =1, n and j != i)
with ith determinant as
reference sequential determinant was then varied from i = 1 to n. The
conformational epitopes is computed.
cutoff distance 6A is found to be optimum.
CE predicts the Conformational
Epitopes with an accuracy of 75%, when
tested using 21 X-ray diffracted co-crystal structure complexes of
antigen and antibodies that are available in PDB database (unpublished).
Scripting language: CGI Perl
Accessibility calculation: Voronoi procedure implemented by McConkey et
Server : Apache on Linux 9.2
IE4+ and Netscape 4 and above.
to Calculate CE
Click on upload button to start
calculation. Once the message "Processing Completed" appears, results
can be accessible for both sequential epitopes and conformational
epitopes using hyperlinks .
User has to upload 3D coordinate
file in PDB format
CE lists both, Conformational
and Sequential epitopes. Results
are saved as html file.
Sample output file
A.S. Kolaskar and
Urmila Kulkarni-Kale (1999). Prediction of three-dimensional structure
and mapping of conformational epitopes of envelope glycoprotein of
Japanese encephalitis virus, Virology, 261, 31-42.
(2003). prediction of 3D structure & function of protein. Ph.D.
Thesis. University of Pune.
A. S. Kolaskar &
Urmila Kulkarni-Kale. CE: An algorithm to predict the conformational
epitopes (in preparation).
Urmila Kulkarni- Kale & A. S.
Prof. A. S.
University of Pune, Pune 411 007
2569 0195/2569 2039
+91 20 2569 0087