PDB数据库在药物化学课程中的应用

通过2个例子说明了PDB数据库在药物化学教学中的应用。这些应用极大地丰富了课堂教学内容,提高了教学质量。     

RepEx: A web server to extract sequence repeats from protein and DNA sequences

Evolution builds up new genetic material from existing ones, not in random, but in highly ordered and eloquent patterns. Most of these sequence repeats are revelatory of valuable information contributing to areas of disease research and function of macromolecules, to name a few. In the age of next generation genome sequencing, rapid and efficient extraction of all unbiased sequence repeats from macromolecules is the need of the hour. In view of this reckoning, an online web-based computing server, RepEx, has been developed to extract and display all possible repeats for DNA and protein sequences. Apart from exact or identical repeats, the server has been designed adeptly to identify and extract degenerate, inverted, everted and mirror repeats from both DNA and protein sequences. The server has striking output displays, featuring interactive graphs and comprehensive output files. In addition, RepEx has been accoutered with an easy-to-use interface and search filters to facilitate a user-defined query or search and is freely available and accessible via the World Wide Web at http://bioserver2.physics.iisc.ac.in/RepEx/.     

Hydrogen Bonds with Fluorine in Ligand–Protein Complexes-the PDB Analysis and Energy Calculations

Fluorine is a common substituent in medicinal chemistry and is found in up to 50% of the most profitable drugs. In this study, a statistical analysis of the nature, geometry, and frequency of hydrogen bonds (HBs) formed between the aromatic and aliphatic C–F groups of small molecules and biological targets found in the Protein Data Bank (PDB) repository was presented. Interaction energies were calculated for those complexes using three different approaches. The obtained results indicated that the interaction energy of F-containing HBs is determined by the donor–acceptor distance and not by the angles. Moreover, no significant relationship between the energies of HBs with fluorine and the donor type was found, implying that fluorine is a weak HB acceptor for all types of HB donors. However, the statistical analysis of the PDB repository revealed that the most populated geometric parameters of HBs did not match the calculated energetic optima. In a nutshell, HBs containing fluorine are forced to form due to the stronger ligand–receptor neighboring interactions, which make fluorine the “donor’s last resort”.     

基于离散小波变换的蛋白质结构、功能与进化关系延时交叉相关分析

基于氨基酸的疏水特性, 将离散小波变换与延时交叉相关分析相结合, 提出了一种分析蛋白质结构、功能和进化关系的新方法——序列小波层次相关法. 以丝氨酸蛋白酶超家族中Trypsin种属为研究对象, 描述了该方法在揭示不同物种的Trypsin蛋白质进化关系中的应用; 以蛋白质酪氨酸磷酸酶、血红蛋白和溶菌酶为例, 采用该方法揭示了蛋白质之间功能与序列的内在关系. 该方法为非参数方法, 具有简单、直观、可视和可操作性强等特点, 且不需要事先知道蛋白质的结构, 仅从蛋白质的氨基酸序列出发即可比较和揭示蛋白质之间的关系.     

Bacterial protein structures reveal phylum dependent divergence

Protein sequence space is vast compared to protein fold space. This raises important questions about how structures adapt to evolutionary changes in protein sequences. A growing trend is to regard protein fold space as a continuum rather than a series of discrete structures. From this perspective, homologous protein structures within the same functional classification should reveal a constant rate of structural drift relative to sequence changes. The clusters of orthologous groups (COG) classification system was used to annotate homologous bacterial protein structures in the Protein Data Bank (PDB). The structures and sequences of proteins within each COG were compared against each other to establish their relatedness. As expected, the analysis demonstrates a sharp structural divergence between the bacterial phyla Firmicutes and Proteobacteria. Additionally, each COG had a distinct sequence/structure relationship, indicating that different evolutionary pressures affect the degree of structural divergence. However, our analysis also shows the relative drift rate between sequence identity and structure divergence remains constant.     

Implementation of Smooth Continuous Camera Trajectories for Viewing PDB and VRML Objects

Summary A parametrically defined camera trajectory enabling the accurate and systematic scanning of the entire surface of virtual (Protein Data Bank (PDB)) proteins is developed and implemented. The smooth continuous path guarantees that each local region of the protein is inspected from a variety of directions in a controlled and uniform manner. Manipulation of several parameters governs the density, character and duration of the scan. Applications to the analysis of other real and virtual objects are also considered.     

The relationship between the sequence identities of alpha helical proteins in the PDB and the molecular similarities of their ligands.

This paper considers the relationship between the percentage sequence identities of protein chains and the molecular similarities of the ligands they bind. Among a set of alpha helical proteins from the PDB, it is found that related proteins tend to bind similar ligands. Furthermore, the property of binding similar ligands can be used to define the categories of "like" and "unlike" pairs of protein chains, separated by an approximate cutoff at a sequence identity of, or somewhat above, 45%. Similarly, the property of binding related protein chains can be used to define "low" and "high" similarity pairs of ligand residues, with a cutoff at a Tanimoto score of 0.70. The ligands bound to two "like" protein chains are five times more likely to be of high similarity than would be expected if protein sequence identity and ligand molecular similarity were independent variables. Nonetheless, the nature of the PDB means that it is unclear whether the same conclusions would be reached with a data set representing an unbiased sample of all protein-ligand complexes in a living cell. The construction of an appropriate data set for such a study represents a significant challenge.     

Internal amino acid sequence analysis of proteins separated by gel electrophoresis after tryptic digestion in polyacrylamide matrix

Summary A method is described for obtaining peptide fragments for sequence analysis from microquantities of proteins separated by 1- or 2-dimensional polyacrylamide gel electrophoresis. After separation by electrophoresis, the proteins were stained with Coomassie Blue and excised. Proteolytic digestion with trypsin was performed directly in the polyacrylamide matrix. The resulting peptide fragments were eluted, separated by reversed phase HPLC, collected and sequenced in a gas phase sequencer. Excellent peptide recoveries allowed generation of extensive internal sequence information from picomole amounts of protein. The method thus overcomes the problem of obtaining amino acid sequence data from N-terminally blocked proteins and provides multiple, independent stretches of sequences that can be used to generate oligonucleotide probes for molecular cloning, to design synthetic peptides for inducing antibodies, and to search sequence databases for related proteins.     

Classification and functional analyses of putative virulence factors of Mycobacterium tuberculosis: A combined sequence and structure based study

The emergence of the drug-resistant mechanisms in Mycobacterium tuberculosis poses the biggest challenges to the current therapeutic measures, which necessitates the identification of new drug targets. The Hypothetical Proteins (HPs), a class of functionally uncharacterized proteins, may provide a new class of undiscovered therapeutic targets. The genome of M. tuberculosis contains 1000 HPs with their sequences were analyzed using a variety of bioinformatics tools and the functional annotations were performed. The functions of 662 HPs were successfully predicted and further classified 483 HPs as enzymes, 141 HPs were predicted to be involved in the diverse cellular mechanisms and 38 HPs may function as transporters and carriers proteins. Furthermore, 28 HPs were predicted to be virulent in nature. Amongst them, the HP P95201, HP P9WM79, HP I6WZ30, HP I6 × 9T8, HP P9WKP3, and HP P9WK89 showed the highest virulence scores. Therefore, these proteins were subjected to extensive structure analyses and dynamics of their conformations were investigated using the principles of molecular dynamics simulations, each for a 150 ns time scale. This study provides a deeper understanding of the undiscovered drug targets and the generated outputs will facilitate the process of drug design and discovery against the infection of M. tuberculosis.     

A statistical analytical approach to predict the secondary structure of proteins from amino acid sequence information

A statistical analytical approach has been used to analyze the secondary structure (SS) of amino acids as a function of the sequence of amino acid residues. We have used 306 non-homologous best-resolved protein structures from the Protein Data Bank for the analysis. A sequence region of 32 amino acids on either side of the residue is considered in order to calculate single amino acid propensities, di-amino acid potentials and tri-amino acid potentials. A weighted sum of predictions obtained using these properties is used to suggest a final prediction method. Our method is as good as the best-known SS prediction methods, is the simplest of all the methods, and uses no homologous sequence/family alignment data, yet gives 72% SS prediction accuracy. Since the method did not use many other factors that may increase the prediction accuracy there is scope to achieve greater accuracy using this approach. Received: 4 May 1998 / Accepted: 17 September 1998 / Published online: 10 December 1998     

Investigation of manganese metal coordination in proteins: a comprehensive PDB analysis and quantum mechanical study

Structural Chemistry - Manganese (Mn) is an important metal that is crucial in biological cell mechanism and function. However, its binding mechanism is poorly characterized. In the present study,...     

Multiple-locus variable number tandem repeats analysis for genetic fingerprinting of pathogenic bacteria

DNA fingerprinting has attracted considerable interest as means for identifying, tracing and preventing the dissemination of infectious agents. Various methods have been developed for typing of pathogenic bacteria, which differ in discriminative power, reproducibility and ease of interpretation. During recent years a typing method, which uses the information provided by whole genome sequencing of bacterial species, has gained increased attention. Short sequence repeat (SSR) motifs are known to undergo frequent variation in the number of repeated units through cellular mechanisms most commonly active during chromosome replication. A class of SSRs, named variable number of tandem repeats (VNTRs), has proven to be a suitable target for assessing genetic polymorphisms within bacterial species. This review attempts to give an overview of bacterial agents where VNTR-based typing, or multiple-locus variant-repeat analysis (MLVA) has been developed for typing purposes, together with addressing advantages and drawbacks associated with the use of tandem repeated DNA motifs as targets for bacterial typing and identification.     

A note on clustering the functionally-related paralogues and orthologues of proteins: a case of the FK506-binding proteins (FKBPs)

The expression patterns of 18 FK506-binding proteins (FKBPs) encoded in the human genome have been established whereas the functional significance of the numerous ORFs coding for FKBP-like sequences remains unknown. Nominal masses of the human FKBPs vary from 12 to 135 kDa. Some large FKBPs consist up to four repeats of the 12 kDa FK506-like binding domain (FKBD) whereas other large FKBPs contain one FKBD linked to different functional domains such as TPRs, leucine-zipper, calmodulin-binding domain etc. The genomes of other eukaryotic organisms, namely D. melanogaster, C. elegans, A. thaliana, S. pombe and S. cerevisiae encode different numbers of the FKBPs' paralogues some of which are orthologues to the human FKBPs. A library of novel algorithms was developed and used for computation of the level of conservation of the hydrophobicity and bulkiness profiles, and the amino acid compositions (AACs) of 247 aligned sequences of FKBPs. The pairwisely-compared hydrophobicity and bulkiness profiles for some combinations of the aligned sequences of the FKBDs yielded high values of the correlation coefficients (CCF). The AACs of some combinations of the aligned sequences of the FKBDs also differed to a low degree. The functionally-related orthologues and paralogues of the FKBPs were clustered by using the following criteria: 1 degrees apparent conservation of the crucial amino acid (AA) residues for peptidylprolyl cis/trans isomerase (PPIase) acitity and binding of some immunosuppressive drugs; 2 degrees convergence of the three mentioned above properties of the polypeptide chain; 3 degrees similarity in the sequence attributes pI and total hydrophobicity index (HI). The clustering method was used for setting up several hypotheses on the emergence of certain classes of the FKBPs in the eukaryotic kingdom.     

嵌入Y型分子筛中钯簇合成与结构的研究

Pd clusters encaged in Y-zeolite (Pd0Y) have been prepared by means of exchanging zeolite HY with [Pd(NH3)4]2+ under microwave radiation. The product formed was deaminized by heating, washed sufficiently with de-ionized water and reduced with hydrogen. The crystal phase diffraction of Pd was not found in the XRD spectrogram of Pd0y. According to polycrystal X-ray diffraction data. Radial Electron Distribution Function (REDF) of Pd0Y was calculated to elucidate the structure of Pd cluster. The results show that the Pd clusters are of the Al type closely packed arrangement. The dimension of them is about 12 Å. They are encaged in the supercage of zeolite Y. Their occupancy on the supercage is as small as 0.06 so that the framework structure of zeolite Y is unchanged. Therefore, the high dispersing Pd cluster aggregating in supercage exhibit strong catalytic effect. The CO-conversion of Pd0Y with Pd 0.72% and 6.13% (in mass fraction) is 67 % and 100 % (in volume fraction) respectively. Evaluation conditions:
mixed gas containing 0.02% CO and air,
space velocity 3000 h-1,
reaction temperature 0 ℃.     

嵌入Y型分子筛中钯簇合成与结构的研究

经微波交换-焙烧-氢还原等过程制备了嵌入Y型分子筛中的钯簇化合物（Pd0Y）应用径向电子分布函数法（RedialElectronDistributionFunction），就其钯原子簇化合物进行了结构的研究．结果说明，嵌入Y型分子筛中的钯原子以A1型密堆积方式排列，聚集成约12大小的原子簇，嵌在Y型分子筛超笼之中．分子筛骨架的作用使族之间分立存在．它在超笼之中的占有率仅为0.06因此Pd0Y化合物既有纳米级金属钯的性能，又有分子筛固有的孔道结构特征．如此含有钯簇的分子筛化合物对一氧化碳完全氧化的反应具有超常的催化活性．     

Estimating structure quality trends in the Protein Data Bank by equivalent resolution

The quality of protein structures obtained by different experimental and ab-initio calculation methods varies considerably. The methods have been evolving over time by improving both experimental designs and computational techniques, and since the primary aim of these developments is the procurement of reliable and high-quality data, better techniques resulted on average in an evolution toward higher quality structures in the Protein Data Bank (PDB). Each method leaves a specific quantitative and qualitative “trace” in the PDB entry. Certain information relevant to one method (e.g. dynamics for NMR) may be lacking for another method. Furthermore, some standard measures of quality for one method cannot be calculated for other experimental methods, e.g. crystal resolution or NMR bundle RMSD. Consequently, structures are classified in the PDB by the method used. Here we introduce a method to estimate a measure of equivalent X-ray resolution (e-resolution), expressed in units of Å, to assess the quality of any type of monomeric, single-chain protein structure, irrespective of the experimental structure determination method. We showed and compared the trends in the quality of structures in the Protein Data Bank over the last two decades for five different experimental techniques, excluding theoretical structure predictions. We observed that as new methods are introduced, they undergo a rapid method development evolution: within several years the e-resolution score becomes similar for structures obtained from the five methods and they improve from initially poor performance to acceptable quality, comparable with previously established methods, the performance of which is essentially stable.     

信号肽对OPMA在枯草杆菌中的分泌表达及其催化活性的影响

将编码枯草杆菌BS168的amyE信号肽(S)的基因与编码多功能淀粉酶OPMA-N和OPMA-G的基因重组, 分别构建了分泌表达多功能淀粉酶OPMA-N和OPMA-G的重组载体pMA5-OPMA-SN和pMA5-OPMA-SG, 并分别在枯草杆菌BS168中实现了OPMA-SN和OPMA-SG的有效分泌表达.SDS-PAGE分析表明, 该信号肽的引入提高了异源蛋白的分泌量, OPMA-SN和OPMA-SG的分泌水平(53%和67%)比OPMA-N和OPMA-G(45%和58%)明显提高, 但此信号肽在分泌表达OPMA-N或OPMA-G后不能被有效切除.活力分析表明, OPMA-SN(5.17 U/mg)和OPMA-N(4.57 U/mg)的活力水平与分泌水平一致, 但OPMA-SG(4.50 U/mg)和OPMA-G(4.65 U/mg)的活力水平却与分泌水平呈反相关性.通过分析OPMA-SN和OPMA-SG分子的空间构象发现, 信号肽的存在不影响OPMA-N的活性部位构象, 但影响OPMA-G的活性部位构象, 从而导致OPOMA-G的催化活性下降.     

The sequence homologies of cytochromes P-450 and active-site geometries

Summary The amino acid sequence alignment of 16 cytochrome P-450 proteins representative of the major families is reported. The sequence matching process has been carried out on the basis of maximum homology by residue type, retention of secondary structure and minimization of deletions/insertions except where additional loop regions exist. From the starting point of known reported sequence homology matching from the literature, a realignment on the basis of conserved residues involved in both structure and function gives rise to a self-consistent set of sequences which correlates with known mechanistic and structural data. Once fitted, these archetypal sequences form a straightforward template for the alignment of all P-450 subfamilies. Computer modelling of the active-site regions constructed from homology with the bacterial form of the enzyme (P-450_CAM) evinces the correct substrate specificity. Furthermore, the construction of the macromolecular assembly of components of the cytochrome P-450 system on the microsomal endoplasmic reticular membrane is presented from the evidence of site-directed mutagenesis, analysis by molecular probes, X-ray crystallography and molecular modelling.     

Systematic investigation of sequence and structural motifs that recognize ATP

Interaction between ATP, a multifunctional and ubiquitous nucleotide, and proteins initializes phosphorylation, polypeptide synthesis and ATP hydrolysis which supplies energy for metabolism. However, current knowledge concerning the mechanisms through which ATP is recognized by proteins is incomplete, scattered, and inaccurate. We systemically investigate sequence and structural motifs of proteins that recognize ATP. We identified three novel motifs and refined the known p-loop and class II aminoacyl-tRNA synthetase motifs. The five motifs define five distinct ATP–protein interaction modes which concern over 5% of known protein structures. We demonstrate that although these motifs share a common GXG tripeptide they recognize ATP through different functional groups. The p-loop motif recognizes ATP through phosphates, class II aminoacyl-tRNA synthetase motif targets adenosine and the other three motifs recognize both phosphates and adenosine. We show that some motifs are shared by different enzyme types. Statistical tests demonstrate that the five sequence motifs are significantly associated with the nucleotide binding proteins. Large-scale test on PDB reveals that about 98% of proteins that include one of the structural motifs are confirmed to bind ATP.