BUDDY-system: A web site for constructing a dataset of protein pairs between ligand-bound and unbound states.

Abstract

Elucidating molecular recognition by proteins, such as in enzyme-substrate and receptor-ligand interactions, is a key to understanding biological phenomena. To delineate these protein interactions, it is important to perform structural bioinformatics studies relevant to molecular recognition. Such studies require a dataset of protein structure pairs between ligand-bound and unbound states. In many studies, the same well-designed and high-quality dataset has been used repeatedly, which has spurred the development of subsequent relevant research. Using previously constructed datasets, researchers are able to fairly compare obtained results with those of other studies; in addition, much effort and time is saved. Therefore, it is important to construct a refined dataset that will appeal to many researchers. However, constructing such datasets is not a trivial task.
We have developed the BUDDY-system, a web site designed to support the building of a dataset comprising pairs of protein structures between ligand-bound and unbound states, which are widely used in various areas associated with molecular recognition. In addition to constructing a dataset, the BUDDY-system also allows the user to search for ligand-bound protein structures by its unbound state or by its ligand; and to search for ligands by a particular receptor protein.
The BUDDY-system receives input from the user as a single entry or a dataset consisting of a list of ligand-bound state protein structures, unbound state protein structures, or ligands and returns to the user a list of protein structure pairs between the ligand-bound and the corresponding unbound states. This web site is designed for researchers who are involved not only in structural bioinformatics but also in experimental studies. The BUDDY-system is freely available on the web.

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TECHNICAL NOTEOpen Access
BUDDY-system: A web site for constructing a
dataset of protein pairs between ligand-bound
and unbound states
Mizuki Morita1,2*, Tohru Terada3, Shugo Nakamura4and Kentaro Shimizu2,4
Abstract
Background: Elucidating molecular recognition by proteins, such as in enzyme-substrate and receptor-ligand
interactions, is a key to understanding biological phenomena. To delineate these protein interactions, it is
important to perform structural bioinformatics studies relevant to molecular recognition. Such studies require a
dataset of protein structure pairs between ligand-bound and unbound states. In many studies, the same well-
designed and high-quality dataset has been used repeatedly, which has spurred the development of subsequent
relevant research. Using previously constructed datasets, researchers are able to fairly compare obtained results
with those of other studies; in addition, much effort and time is saved. Therefore, it is important to construct a
refined dataset that will appeal to many researchers. However, constructing such datasets is not a trivial task.
Findings: We have developed the BUDDY-system, a web site designed to support the building of a dataset
comprising pairs of protein structures between ligand-bound and unbound states, which are widely used in
various areas associated with molecular recognition. In addition to constructing a dataset, the BUDDY-system also
allows the user to search for ligand-bound protein structures by its unbound state or by its ligand; and to search
for ligands by a particular receptor protein.
Conclusions: The BUDDY-system receives input from the user as a single entry or a dataset consisting of a list of
ligand-bound state protein structures, unbound state protein structures, or ligands and returns to the user a list of
protein structure pairs between the ligand-bound and the corresponding unbound states. This web site is
designed for researchers who are involved not only in structural bioinformatics but also in experimental studies.
The BUDDY-system is freely available on the web.
Findings
Elucidating molecular recognition by proteins is one of
the keys to understanding biological phenomena. Struc-
tural bioinformatics studies relevant to molecular recog-
nition, such as analysis of conformational changes upon
ligand binding [1-4], development of methods for pre-
dicting ligand binding sites [5-7], and development of
molecular docking tools [8-10], require a dataset of pro-
tein structure pairs between ligand-bound and unbound
states (Figure 1).
The BUDDY-system features a flexible definition of a
ligand and allows the user to change various options via
its web interface. The BUDDY-system is based on a pre-
mise that differs from existing structural bioinformatics
systems in terms of what is considered a ligand. In pre-
vious studies, a ligand was defined as all heterogeneous
(HETATM) molecules in the Protein Data Bank (PDB)
[11] files [1,12], all HETATM molecules except for low-
molecular ions (e.g., Zn2+, Mn2+, PO43-, and SO42-) [4],
or HETATM clusters forming many inter-atomic con-
tacts with protein atoms [13]. This variety of ligand defi-
nitions implies that it is very difficult to specifically
define a ligand. Here, we define a ligand as molecules
that can dissociate from proteins; consequently, a cer-
tain protein can be found with a ligand in some entries
in PDB and without it in other entries. Under this defi-
nition, a ligand is not determined specifically but instead
depends on each pair of PDB entries. For example, the
* Correspondence: mizuki@bi.a.u-tokyo.ac.jp
1Department of Fundamental Research, National Institute of Biomedical
Innovation (NIBIO), 7-6-8 Saito Asagi, Ibaraki, Osaka 567-0085, Japan
Full list of author information is available at the end of the article
Morita et al. BMC Research Notes 2011, 4:143
http://www.biomedcentral.com/1756-0500/4/143
© 2011 Morita et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons
Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in
any medium, provided the original work is properly cited.

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structure of fructose-1,6-bisphosphatase (F16BPase) [14],
which catalyzes the hydrolysis of d-fructose 1,6-bispho-
sphate (FBP) to d-fructose 6-phosphate (F6P) and phos-
phate (Pi), has been demonstrated several times in
different binding states (Figure 2): F16BPase in free
form ([PDB:2FBP]); with F6P in the active site
([PDB:1RDX]); with F6P and adenosine monophosphate
(AMP) in the allosteric site ([PDB:1FBP]); and with F6P,
AMP, and the anilinoquinazoline inhibitor (PFE) in the
non-native allosteric site ([PDB:1KZ8]). If a ligand is
defined specifically as “HETATM molecules except for
low-molecular ions,” [PDB:2FBP] would be reported as
existing in the ligand-unbound state and all the others
intheligand-bound state.
[PDB:1RDX] exists in the ligand-bound state against
[PDB:2FBP], it also exists in the ligand-unbound state
against [PDB:1FBP] and [PDB:1KZ8]. Likewise, while
[PDB:1FBP] is in the ligand-bound state against
[PDB:2FBP] and [PDB:1RDX], it is also present in a
ligand-unbound state against [PDB:1KZ8]. The flexible
ligand definition in the BUDDY-system enables the user
to obtain all possible ligand-bound and unbound state
pairs of F16BPase.
We plan to implement more advanced search options
in the future, such as protein sequence similarity search
and chemical structure search from SMILES.
However,although
Methods
The procedure for constructing a dataset of protein
pairs between ligand-bound and unbound states (called
bound/unbound-pairs) in the BUDDY-system consists of
the following 3 steps: (1) finding all pairs of the same
proteins or homologues in all the PDB entries to pre-
pare an initial dataset, (2) screening bound/unbound-
pairs from the initial dataset to prepare a super dataset,
and (3) finding suitable pairs for the user’s request from
the super dataset after the user submits a request
(Figure 3). The first 2 steps are carried out in advance,
and the third step can be achieved after the user enters
input data. The details are as follows. (1) The BUDDY-
system finds pairs of the same proteins or homologues
from all of the PDB entries based on their sequence
identity to prepare an initial dataset (the sequence iden-
tity threshold can be specified by the user via the web
interface). Here, a chain shorter than N amino acids is
defined as “a peptide chain” and is considered a ligand
(N can be specified by the user via the web interface).
This option is useful, especially when a protein has
short amino acid chains that are essential for its func-
tion (e.g., insulin). (2) Next, the BUDDY-system screens
the bound/unbound-pairs to prepare a super dataset
from the initial dataset. Initially, the BUDDY-system
compiles HETATM lists of both PDB entries in a pair,
respectively. Here, when an HETATM molecule appears
more than once in a PDB entry, it is listed only once in
its HETATM list. Furthermore, HETATM molecules
Figure 1 Example of a protein structure pair between ligand-
bound and unbound states. (a) A ligand-bound state structure of
6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase (HPPK)
([PDB:1HQ2]) and (b) an unbound state structure ([PDB:1IM6]). The
ligand is represented by dark spheres. The BUDDY-system allows
users to search for this type of pair by its ligand as a search query,
bound states and ligands by its unbound state, and unbound states
from its bound state. The user can input 1 query or a set of such
queries as a dataset.
Figure 2 Examples illustrating the difficulty in defining a
ligand. F16BPase tetramer (a) in free form ([PDB:2FBP]), (b) with F6P
(red sphere) in the active site ([PDB:1RDX]), (c) with F6P and AMP
(yellow sphere) in the allosteric site ([PDB:1FBP]), and (d) with F6P,
AMP, and PFE (green sphere) in the non-native allosteric site
([PDB:1KZ8]). Although [PDB:1RDX] is in a ligand-bound state against
[PDB:2FBP], it is also in a ligand-unbound states against [PDB:1FBP].
Further, although [PDB:1FBP] is in a ligand-bound state against
[PDB:2FBP] and [PDB:1RDX], it is also in a ligand-unbound state
against [PDB:1KZ8]. If a ligand is defined specifically as “HETATM
molecules except for low-molecular ions,” all entries but [PDB:2FBP]
are obtained in a ligand-bound state. The flexible ligand definition
in the BUDDY-system enables the user to obtain all possible ligand-
bound and unbound states pairs of F16BPase.
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that are defined as “not considered as a ligand” will be
excluded from the HETATM list. If the PDB file has
chains shorter than N amino acids in the ATOM record
(N can be decided by the user via the web interface),
they are considered “peptide chains.” The BUDDY-sys-
tem then compares 2 HETATM lists and peptide chains
from 2 PDB entries in a pair and judges whether this
pair is a bound/unbound-pair in the following manner:
(2-i) when the contents of 2 HETATM lists and peptide
chains are identical, this pair is not regarded as a
bound/unbound-pair; and (2-ii) when those are not
identical, a pair is a bound/unbound-pair if 1 HETATM
list is included in another list. (3) Finally, after the user
inputs ligand-bound state protein structures, unbound
state protein structures, or ligands into the BUDDY-sys-
tem, bound/unbound-pairs that fit the user’s request are
selected from the super dataset and are returned to the
user. The user can (3-i) upload their own datasets
including a PDB ID list of ligand-bound state protein
structures, unbound state protein structures, or a
HETATM ID list of ligands; (3-ii) choose one of the
readymade datasets of ligand-bound state protein struc-
tures, such as BindingDB [15] and PLD [16] (whose use
of each has been generously permitted by the authors
listed in references 14 and 15, respectively); or (3-iii)
input one PDB ID of a ligand-bound state protein struc-
ture or an unbound state protein structure, or one
HETATM ID of a ligand. The file formats of the input
and output datasets are described on the BUDDY-
system website. The parameters that the user can select
are the cut-off value of X-ray resolution, the sequence
identity when making a bound/unbound-pair, and the
definition of a peptide chain.
Example Usage
Here, we show examples of using the BUDDY-system.
Table 1 shows the results obtained from the BUDDY-
system when a list of PDB entries of ligand-bound state
proteins, which were obtained from various databases or
datasets available on the Internet, were input with the
following default parameters: X-ray resolutions equal to
or better than 2.5 Å were allowed, a sequence identity
between ligand-bound and unbound state protein equal
to 100% was used, and chains shorter than 30 amino
acids were considered peptide ligands. In the example
shown in Table 1, when PDB entries obtained from Bin-
dingDB were input, at least 1 corresponding unbound
state entry was found for 484 of 1,485 input ligand-
bound state protein entries, and the number of total
pairs was 4,629. Interestingly, at least 1 unbound state
PDB entry was found for approximately 30% of the
input ligand-bound state protein structures for any of
the datasets in Table 1. Additionally, a large portion of
these ligand-bound state structures was paired with only
1 corresponding unbound state protein structure.
Although this number of returned pairs would increase
or decrease depending on the parameters used, these
examples in Table 1 demonstrate that a dataset of
Figure 3 Schematic diagram illustrating the construction of a
dataset in the BUDDY-system. The process of constructing a pair
dataset consists of the following 3 steps: (1) all pairs of the same
proteins or homologues are obtained from the entire PDB entries to
prepare an initial dataset, (2) protein structure pairs of ligand-bound
and unbound states are screened from the initial dataset to prepare
a super dataset, and (3) the pairs that fit the user’s request are
selected from the super dataset after the user submits a request.
Table 1 Summary of the results obtained using the
BUDDY-system against various datasets
Input dataseta
Versionb
The number of protein
structures
Inputc
Output Ad
Output Be
AffinDB [17]
BindingDB [15]
PDBbind [18]
PLD [16]
CCDC/Astex Test Set [8]
Astex Diverse Set [10]
Non-redundantf
2008-03-17
2008-03-17
v2007
v1.3
-
-
-
476
1,485
3,124
485
305
85
4,544
157
484
996
137
91
25
1,172
2,257
4,629
7,000
1,277
611
170
7,901
This table shows the results obtained from the BUDDY-system when various
databases were input as lists of ligand-bound states proteins. Taking
BindingDB as an example, when 1,485 PDB entries obtained from BindingDB
were input as a ligand-bound state, at least 1 corresponding unbound state
PDB entry was found for 484 of 1,485 input entries; a total of 4,629 pairs were
obtained.
aEach input dataset contains a list of PDB entries of ligand-bound state
proteins.
bThe version of the input dataset or the date when the input dataset was
downloaded.
cThe number of ligand-bound state protein structures in each input dataset.
dThe number of unique proteins that were paired with at least one ligand-
unbound state entry by the BUDDY-system.
eThe number of total pairs.
fUnion of all ligand-bound state protein datasets.
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bound/unbound-pairs can be readily obtained with the
BUDDY-system. The datasets obtained here are essential
for elucidating molecular recognition by proteins in stu-
dies that investigate conformational changes involved in
enzyme reactions, developments of ligand binding site
prediction, and components involved in molecular dock-
ing. The BUDDY-system is the first web site that the
authors are aware of that supports the construction of
such a dataset according to the user’s input dataset and
parameters. In addition, because the ligand is allowed a
more flexible definition, this web server is useful to
exhaustively search for ligands or ligand-bound and
unbound state structures that are of interest to the user.
Availability and Requirements
The BUDDY-system is freely available at URL http://
www.bi.a.u-tokyo.ac.jp/services/buddy/.
Abbreviations
AMP: Adenosine monophosphate; F16BPase: Fructose-1,6-bisphosphatase;
F6P: d-fructose 6-phosphate; HPPK: 6-Hydroxymethyl-7,8-dihydropterin
pyrophosphokinase; PDB: Protein Data Bank; Pi: Phosphate.
Acknowledgements
The authors thank Dr. John Mitchell and Dr. Michael Gilson for granting
permission to use their datasets. The authors also thank Dr. Kazuya
Sumikoshi for technical contributions. This work was partially supported by
Grant-in-Aid for Young Scientists (B) and Grant-in-Aid for Scientific Research
on Priority Areas Systems Genomics from the Ministry of Education, Culture,
Sports, Science and Technology of Japan.
Author details
1Department of Fundamental Research, National Institute of Biomedical
Innovation (NIBIO), 7-6-8 Saito Asagi, Ibaraki, Osaka 567-0085, Japan.
2Institute for Bioinformatics Research and Development (BIRD), Japan
Science and Technology Agency (JST), 4-1-8 Honcho, Kawaguchi, Saitama
332-0012, Japan.3Agricultural Bioinformatics Research Unit, The University of
Tokyo, 1-1-1 Yayoi, Bunkyo, Tokyo 113-8657, Japan.4Department of
Biotechnology, The University of Tokyo, 1-1-1 Yayoi, Bunkyo, Tokyo 113-8657,
Japan.
Authors’ contributions
MM developed the concept and designed the algorithm and its
implementation. TT provided valuable suggestions on the manuscript. SN
contributed to the implementation of the web site. KS reviewed and tested
the software. All authors read and approved the final version of the
manuscript.
Competing interests
The authors declare that they have no competing interests.
Received: 21 December 2010 Accepted: 22 May 2011
Published: 22 May 2011
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doi:10.1186/1756-0500-4-143
Cite this article as: Morita et al.: BUDDY-system: A web site for
constructing a dataset of protein pairs between ligand-bound and
unbound states. BMC Research Notes 2011 4:143.
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