Download Sequence and Structural Analysis of Ligand Binding Sites in

Sequence and Structural Analysis of Ligand Binding Sites
in Membrane Proteins
M. Xavier Suresh
[email protected]
M. Michael Gromiha
[email protected]
M. Suwa
[email protected]
Computational Biology Research Center (CBRC), AIST, 2-42 Aomi, Koto-ku, Tokyo 135-0064, Japan
Keywords: membrane proteins, database analysis, ligand interaction
1
Introduction
Known to be involved in many functionally important activities such as signal transduction, ion transport,
and cell-cell communications, membrane proteins are an important class of proteins. In recent years the
numbers of entries are gradually increasing in the structural databases. The sequence and structural analysis
on the interactions between the membrane proteins and ligands would help to derive structural principles in
protein-ligand complexes which provides deep insights in to the mechanism of protein-ligand interactions
and their function. Therefore we have analyzed the membrane protein-ligand complexes through contacts
between protein and ligand, accessible surface area, secondary structure, and location of the residues, etc.
Atomic level analyses are generally applicable to the identification of surface epitopes and binding pockets
for development of diagnostic reagents, therapeutics and vaccines.
2
Methods
2.1 Dataset
Table 1: Dataset.
The protein structures used in this present study have been obtained
Group
No. of complexes
from PDBTM [1], a database of transmembrane proteins derived Metal/ion binding
134
from the protein data bank (PDB). Atomic coordinates were Sugar binding
42
obtained from the PDB [2]. Of the 521 transmembrane protein Others
140
structures, excluding the NMR structures, fragments and repeated
structures, 173 are found to be complexed with various ligands. Based on the bound ligands they are further
classified into different groups (Table 1). These groups contain both α-helical and β-barrel proteins as well.
Atom contacts and residue contacts were calculated as follows. An atom in a protein is said to be in
contact if the distance between this atom and any one of the atom in the ligand is within a distance cutoff 4.5
Å. Similarly an amino acid residue within a protein sequence was designated as a binding site residue if its
side chain or backbone atoms fell within a cutoff distance of 4.5 Å from any atom within a binding ligand.
Secondary structure and solvent accessibility or accessible surface area (ASA) values of the selected
membrane protein-ligand complexes in the dataset have been calculated using DSSP program [3]. Absolute
values of ASA, thus obtained are normalized to relative values as described in Ahmad and Gromiha [4]. The
frequency of occurrence of the dipeptide together with the interacting residues was also analyzed.
3
Results and Discussion
3.1 Atom Contacts and Residue Contacts
The atom contact analysis on 173 membrane protein-ligand complexes shows that 7.23%, 27.91% and
64.86% of atom contacts are around the ligands at the vicinity of 0-3.0, 3.0-4.5 and 4.5-6.0 Å which reveals
the fact that, in addition to the direct interactions, the influence of the neighboring atoms and residues may
also play key roles in the binding of these ligands at the binding site [5]. The binding frequencies of 20
amino acid residues in the three different groups are depicted in Fig. 1. In metal binding complexes, the
preference of Cys is significant. 5% of Cys in disulphide bridges whereas 3% involved both in disulphide
and residue contacts. At an average 2 disulfide bridges were found in 27 complexes. Arg is significant in
sugar binding complexes whereas Trp is significant in other complexes because of large ligands (eg.
photosynthetic reaction center, chlorophyll). In order to enhance our understanding, further the contacts were
analyzed at the main chain and side
chain level. The main chain contact
of Cys in metals (>13%) is
significant whereas the in side chain
contacts, His in metals, Arg in sugar
binding complexes, and Trp in other
complexes are observed more may be
because of the bulkiness of the side
chains.
3.2 Accessible Surface Area and
Secondary Structure
Accessible surface area analysis
shows that most of the residues in
contact have ASA in the range 0-20%,
Figure 1: Binding frequencies of 20 amino acids.
indicates that the interacting residues
(more than 70%) are buried (for α-helical proteins 75% and β-barrel proteins 60%) whereas a fraction of
residues are exposed. The pattern observed for the relationship between the frequency of residues at various
ranges of ASA is similar to that obtained by Bartlett et al. [6] upon analyzing the active sites of enzyme
protein structures. The threshold of exposure is 25% in enzyme proteins, but in membrane proteins it is 40%.
This indicates that in membrane proteins the binding site residues are not completely buried. In conclusion,
membrane proteins have binding sites with buried and partially exposed residues.
Analysis of secondary structural preference reveals that the overall preference of the residues in contact
are towards α-helices whereas in α-helical proteins the preference is helix (73%) as expected. In β-barrel
proteins 70% of the residues in contact prefers the β-sheet conformation. About 20% of the residues prefers
random coil or loops. Helix is more preferred for sugar binding complexes when compared to sheet and coil.
3.3 Neighboring Residue Information
We have also analyzed the dipeptide information. Analysis shows that 8.71% of dipeptides are in contact
while it is 13.43% for single amino acid contact. Only 3.25% dipeptides occur more than 50%. Analysis of
the neighboring residues indicates that the role of Cys is predominant. Cys in N-/C- termini is more preferred.
The occurrence of His and Met are more in highly preferred dipeptides, which dictates that these residues
may have great affinity for ligand binding in membrane protein-ligand complexes.
4
Conclusions
Our results are expected to provide a better understanding of the molecular mechanisms involved in ligand
binding and form basis for predicting binding site residues in membrane proteins. Further it can also aid in
identifying potential ligand binding sites. The suggested role of the neighboring residues is also important for
the ligand binding. It is hoped that this information will effectively be used function annotation and aid in
drug design initiatives.
Acknowledgments
MXS is a Post Doctoral Fellow of JSPS, Japan.
References
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