integral membrane proteins represent about 2030% of the total proteins of various organisms ( 13 ) . despite their important physiological roles , they are highly underrepresented in the protein structure database ( pdb ) ( 4 ) , due to the difficulties in crystallizing them in an aqueous environment . among the > 25  000 structures that are deposited in the pdb so far , the number of membrane protein structures hardly exceeds 400 , which represents around 3040 different folds only .
a complete list of transmembrane proteins ( tmps ) is difficult to extract from the pdb that is dominated by water - soluble proteins , mainly because the annotation of pdb files is not reliable in terms of the transmembrane character of the deposited protein structures .
transmembrane into the search field on the pdb home page , the search resulted in 1508 hits ( in july , 2004 ) .
there are only 310 of these proteins , which are really tmps , whereas many others contain only the water - soluble domain of tmps . at the same time
therefore , to determine the type of a protein in the pdb , the actual coordinates of the proteins have to be analyzed . a separate issue that is specific to tmp structures is the identification of membrane - spanning regions .
this information is not included in the pdb files , as the lipid molecules are removed before the process of structure determination .
therefore , no direct experimental data is available about how the protein spans the membrane ( 5 ) .
although the transmembrane character and the likely position of the membrane can be guessed by inspecting individual cases to ensure completeness and consistency , an automated approach is required to identify membrane - spanning regions .
the structures of these proteins are often of low resolution , or correspond to only very small fragments , in which cases automatic assignment can be quite challenging .
we have recently developed an algorithm ( 6 ) , called tmdet , to find the most likely position of the membrane , and to distinguish between transmembrane and non - transmembrane proteins or protein segments by using their coordinates only .
the tmdet algorithm allows the appropriate classification of low - resolution structures , as well as cases when only a fragment or a partial structure of a multichain protein complex is available .
the discrimination power of tmdet algorithm is > 98% ; therefore , it can be used to create and maintain a database that contains the tmps of known 3d structures , as well as to calculate the possible membrane localization of these proteins . here
we present the details of the resulting database , pdb_tm , and its usage on the world wide web .
first , it assigns a transmembrane character for each entry in the pdb , which allows the construction of a comprehensive and up - to - date list of all membrane proteins and segments with known structures .
second , it identifies the location of the lipid bilayer that is relative to the coordinates system of the molecule .
for example , it can help to assign whether a binding site is located in the lipid or in the aqueous phase , which can be important to design a drug that binds to a certain part of a receptor .
what is more , since pdb_tm identifies all tmps in the pdb , the rest of the pdb is a pure database of non - transmembrane proteins , therefore such information might be useful for scientists who work on databases of water - soluble proteins .
the technical details of the tmdet algorithm , as well as the initial version of pdb_tm database can be found in our recent article ( 6 ) . here
we briefly summarize the creation of the database : tmdet algorithm was applied to all files in the pdb . for the sake of simplicity , virus and
pilus proteins as well as entries that contained nucleotides ( rna or dna sequences ) and peptides < 15 residues in length were omitted .
the tmdet algorithm is based on an empirical measure , the q - value ( 6 ) , which describes the manner in which a given slice of a protein fits into a membranous environment . for each protein ,
an exhaustive search is carried out by varying the position and orientation of the slice , in order to find the maximum of the q - values with the corresponding most likely location of the membrane .
if the tmdet algorithm resulted in a maximum q - value below a predefined threshold , the pdb entry was classified as a globular protein .
if the value was above an upper selection limit , the pdb entry was classified as a tmp . in a small number of cases
, for which the q - values fell between the lower and upper selection limit , the transmembrane character could not be assigned confidently . in theses cases ,
the decision was made manually based on the annotation of the pdb file , the annotation of the corresponding swiss - prot entries ( 7 ) , and when in doubt by checking the literature . for all identified tmps , our algorithm also determined the number and type of transmembrane segments , localization of membrane that is relative to the protein coordinates and localization of each sequence part that is relative to the membrane .
the number of transmembrane segments does not include non - canonical elements within the membrane region .
some alpha helical proteins contain segments that do not cross the membrane , but turn back at the middle of the membrane [ e.g. aquaporin , 1fqy , ( 8) ] , whereas the inside of the pore that is formed by beta - barrel proteins are basically shielded from the membrane , and can accommodate alpha helical segments as well [ e.g. the translocator domain of a bacterial autotransporter , 1uyo , ( 9 ) ] .
the algorithm is able to identify the correct topology even when these elements are present in the structure .
the database is updated every week when the new pdb entries are released , by running the tmdet algorithm on each new pdb file
. a typical run on an average pdb file takes a few seconds on a pentium 4 , 2.4 ghz personal computer , thus the update of the database is around one or two hours for the computer plus some minutes for checking the results by a human expert .
the pdb_tm database can be regarded as an extension of the pdb database , because it contains additional information for each pdb entry .
therefore , the database is organized similarly to the pdb , the entries are identified by their pdb code and are grouped in subdirectories according to the middle two characters of their codes .
the properties of proteins determined by the tmdet algorithm are described in xml format , and the corresponding document type definition can be found on the pdb_tm homepage . for non - transmembrane proteins , the xml files contain only the information that the protein is not of the transmembrane type , whereas for transmembrane proteins , the xml files contain all the information that were gathered and calculated by the tmdet algorithm .
these are the q - value on which the selection was made ; the type of the protein according to the secondary structure of its membrane - spanning residues that were determined by the dssp algorithm ( alpha - helical , beta - barrel or unstructured ) ( 10 ) ; and some additional information on keyword search in the corresponding swiss - prot and pdb files .
it is given by the transformation matrix , which rotates the molecule such that the normal of the membrane plane is parallel with the z - axis and translates it , placing the new origin along this axis at the membrane half - width .
the start and end position of the normal vector of the membrane plane is also specified .
the length of the normal vector is equal to the half - width of the membrane .
the last part of the files contains the localization of each sequence parts that is relative to the membrane for each chain that builds the biological active molecule .
because the inside and outside orientation of proteins can not be determined from its coordinates , we use side - one and side - two notation to distinguish between the two sides of the membrane .
the file contains information for all protein chains included in the biological oligomer structure ; therefore , the number of chains can be different in a pdb_tm entry and in the corresponding pdb file .
the definition and the calculation of biological active molecule are described in our previous article .
there are various ways to find information on a protein in the pdb_tm database : the user can find it by entering the protein pdb code , by searching with keywords in the pdb or by entering some properties of the tmp , such as its type or the number of transmembrane segments .
if the search resulted in multiple hits , the list of hits is displayed , and the wanted molecule can be easily selected from the list .
figure 1 shows the explore window of the aquaporin molecule ( pdb code : 1fqy ) , which is a graphical representation of the corresponding xml file .
the database is downloadable , either the full database in one big file , or its subsets containing the alpha helical or the beta - barrel tmps .
, two typical pictures can be viewed on the visualization window , which is made by the pymol molecular graphics software ( 11 ) .
the molecule in this picture is colored according to the identified localization of the segments , which discriminates between transmembrane regions , membrane loops , inner membrane segments as well as the two sides of the water - soluble regions .
currently the pdb_tm database is updated semi - automatically every week right after the weekly update of the pdb .
there is a family classification on the home page representing the situation as of early 2004 .
work is in progress that will allow the automated update of the family classification as well .
while currently tmps can be viewed on pictures made by the pymol molecular graphic software , we plan to extend the range of visualization tools to provide rasmol script , pymol script or vrml , similar to the pdb site .
this work was sponsored by grants bio-0005/2001 , otka t34131 , d42207 and f043609 . z.d . and
 pdb_tm is a database for transmembrane proteins with known structures . 
 it aims to collect all transmembrane proteins that are deposited in the protein structure database ( pdb ) and to determine their membrane - spanning regions . 
 these assignments are based on the tmdet algorithm , which uses only structural information to locate the most likely position of the lipid bilayer and to distinguish between transmembrane and globular proteins . 
 this algorithm was applied to all pdb entries and the results were collected in the pdb_tm database . by using tmdet algorithm , the pdb_tm database can be automatically updated every week , keeping it synchronized with the latest pdb updates . the pdb_tm database is available at http://www.enzim.hu/pdb_tm . 
INTRODUCTION
DATABASE CREATION AND MAINTENANCE
DATABASE ORGANIZATION, FILE FORMATS AND ANNOTATIONS
DATA ACCESS AND VISUALIZATION
FUTURE DIRECTION
ACKNOWLEDGEMENTS