Comparative modeling of a GABAA alpha1 receptor using three crystal structures as templates

We built a model of a GABAA alpha1 receptor (GABAAR) that combines the ligand binding (LBD) and the transmembrane domains (TMD). We used six steps: (1) a four-alpha helical bundle in the crystal structure of bovine cytochrome c oxidase (2OCC) was identified as a template for the TMD of a single subunit. (2) The five pore-forming alpha helices of a bacterial mechanosensitive channel (1MSL) served as a template for the pentameric ion channel. (3) Five copies of the tetrameric template from 2OCC were superimposed on 1MSL to produce a homopentamer containing 20 alpha helices arranged around a funnel-shaped central pore. (4) Five copies of the GABAAR sequence were threaded onto the alpha-helical segments of this template and inter-helical loops were generated to produce the TMD model. (5) A model of the LBD was built by threading the aligned sequence of GABAAR onto the crystal structure of the acetylcholine binding protein (1I9B). (6) The models of the LBD and the TMD were aligned along a common five-fold axis, moved together along that axis until in vdW contact, merged, and then optimized with restrained molecular dynamics. Our model corresponds closely with recently published coordinates of the acetylcholine receptor (1OED) but also explains additional features. Our model reveals structures of loops that were not visible in the cryoelectron micrograph and satisfies most labeling and mutagenesis data. It also suggests mechanisms for ligand binding transduction, ion selectivity, and anesthetic binding.     

Homology modeling of four Y-family, lesion-bypass DNA polymerases: the case that E. coli Pol IV and human Pol kappa are orthologs, and E. coli Pol V and human Pol eta are orthologs

Y-family DNA polymerases (DNAPs) are a superfamily of evolutionarily related proteins that exist in cells to bypass DNA damage caused by both radiation and chemicals. Cells have multiple Y-family DNAPs, presumably to conduct translesion synthesis (TLS) on DNA lesions of varying structure and conformation. The potent, ubiquitous environmental mutagen/carcinogen benzo[a]pyrene (B[a]P) induces all classes of mutations with G-->T base substitutions predominating. We recently showed that a G-->T mutagenesis pathway for the major adduct of B[a]P ([+ta]-B[a]P-N2-dG) in Escherichia coli depends on Y-family member DNAP V. Since no X-ray crystal study for DNAP V has been reported, no structure is available to help in understanding the structural basis for dATP insertion associated with G-->T mutations from [+ta]-B[a]P-N2-dG. Herein, we do homology modeling to construct a model for UmuC, which is the polymerase subunit of DNAP V. The sequences of eight Y-family DNAPs were aligned based on the positioning of conserved amino acids and an analysis of conserved predicted secondary structure, as well as insights gained from published X-ray structures of five Y-family members. Starting coordinates for UmuC were generated from the backbone coordinates for the Y-family polymerase Dpo4 for reasons discussed, and were refined using molecular dynamics with CHARMM 27. A survey of the literature revealed that E. coli DNAP V and human DNAP eta show a similar pattern of dNTP insertion opposite a variety of DNA lesions. Furthermore, E. coli DNAP IV and human DNAP kappa show a similar dNTP insertional pattern with these same DNA lesions, although the insertional pattern for DNAP IV/kappa differs from the pattern for DNAPs V/eta. These comparisons prompted us to construct and refine models for E. coli DNAP IV and human DNAPs eta and kappa as well. The dNTP/template binding pocket of all four DNAPs was inspected, focusing on the array of seven amino acids that contact the base of the incoming dNTP, as well as the template base. DNAPs V and eta show similarities in this array, and DNAPs IV and kappa also show similarities, although the arrays are different for the two pairs of DNAPs. Thus, there is a correlation between structural similarities and insertional similarities for the pairs DNAPs V/eta and DNAPs IV/kappa. Although the significance of this correlation remains to be elucidated, these observations point the way for future experimental studies.     

Comparative protein modeling of methionine S-adenosyltransferase (MAT) enzyme from Mycobacterium tuberculosis: a potential target for antituberculosis drug discovery

Mycobacterium tuberculosis (Mtb) is a successful pathogen that overcomes the numerous challenges presented by the immune system of the host. In the last 40 years few anti-TB drugs have been developed, while the drug-resistance problem is increasing; there is thus a pressing need to develop new anti-TB drugs active against both the acute and chronic growth phases of the mycobacterium. Methionine S-adenosyltransferase (MAT) is an enzyme involved in the synthesis of S-adenosylmethionine (SAM), a methyl donor essential for mycolipid biosynthesis. As an anti-TB drug target, Mtb-MAT has been well validated. A homology model of MAT has been constructed using the X-ray structures of E. coli MAT (PDB code: 1MXA) and rat MAT (PDB code: 1QM4) as templates, by comparative protein modeling principles. The resulting model has the correct stereochemistry as gauged from the Ramachandran plot and good three-dimensional (3D) structure compatibility as assessed by the Profiles-3D score. The structurally and functionally important residues (active site) of Mtb-MAT have been identified using the E. coli and rat MAT crystal structures and the reported point mutation data. The homology model conserves the topological and active site features of the MAT family of proteins. The differences in the molecular electrostatic potentials (MEP) of Mtb and human MAT provide evidences that selective and specific Mtb-MAT inhibitors can be designed using the homology model, by the structure-based drug design approaches.     

Comparative pharmacophore development for inhibitors of human and rat 5-alpha-reductase

There are a number of diseases where the 5-alpha-reductase (5AR) enzyme is of therapeutic interest as a drug target. Currently the crystal structure for 5-alpha-reductase is unavailable, thus ligand-based pharmacophore techniques are beneficial in the drug development process. We have developed pharmacophores to aid inhibitor design for both human types I (preliminary) and II 5-alpha-reductase isozymes and also the rat type II isozyme. To our knowledge, these are the first published pharmacophores for inhibitors of the human type I and rat type II enzymes. A comparison between isozymes and the previously published human type II isozyme pharmacophore is also presented.     

The whey acidic protein family: a new signature motif and three-dimensional structure by comparative modeling

Whey acidic proteins (WAP) from the mouse, rat, rabbit, camel, and pig comprise two "four-disulfide core" domains. From a detailed analysis of all sequences containing this domain, we propose a new PROSITE motif ([KRHGVLN]-X-?PF?-X-[CF]-[PQSVLI]-X(9,19)-C-?P?-X-[DN]-X-?N? -[CE]-X(5)-C-C) to accurately identify new four-disulfide core proteins. A consensus model for the WAP proteins is proposed, based on the human mucous proteinase inhibitor crystal structure. This article presents a detailed atomic model for the two-domain porcine WAP sequence by comparative modeling. Surface electrostatic potential calculations indicate that the second domain of the pig WAP model is similar to the functional human mucous proteinase inhibitor domains, whereas the first domain may be nonfunctional.     

Differences in active-site microarchitecture explain the dissimilar behaviors of PBP5 and 6 in Escherichia coli

Out of the four DD-carboxypeptidases (DD-CPases) in Escherichia coli, only penicillin-binding protein (PBP) 5 performs physiological functions such as maintaining cell shape; its nearest homolog, PBP6, cannot perform such functions. Moreover, unlike PBP6, PBP5 efficiently processes both beta-lactam, and peptide substrates. The crystal structure of PBP5 reveals strong inter-residue hydrogen-bonding interactions around the active site, which favor its catalytic activity. However, the recently solved crystal structure of PBP6 cannot explain the reason for the observed functional discrepancies between the two proteins. Enzymatic analyses indicate that moving the morphology maintenance domain from one protein to another can alter the affinities and activities of PBP5 and 6 toward their substrates. To determine why the activities of these enzymes differ, we used molecular modeling, and docking analyses with substrate-mimetic ligands to estimate how amino-acid alterations in the morphology maintenance domain would affect the structure of PBP and hence its substrate specificity. The results obtained from kinetic analyses were directly correlated to the three-dimensional structures of the PBPs determined through in silico analyses, indicating a change in the active-site microarchitectures of PBP5 and 6 as a plausible cause of the difference in their biochemical behaviors.     

Comparison of homology models and X-ray structures of the nuclear receptor CAR: assessing the structural basis of constitutive activity

The constitutive androstane receptor (CAR) possesses an intrinsic basal activity whose structural basis has been analysed during the last decade. Recently, we published a homology model of the CAR ligand binding domain (LBD) based on the X-ray structures of the closely related pregnane X (PXR) and vitamin D (VDR) receptor. A detailed analysis of the homology model and molecular dynamics (MD) simulations afforded us to propose a potential mechanism underlying the constitutive activity of CAR. Almost simultaneously, X-ray structures of human and mouse CAR LBD were released. In the present study, a detailed analysis and comparison of homology model and X-ray structures is carried out in order to evaluate the quality and reliability of our homology modelling procedure. The hypothesis of the constitutive activity which we proposed on the basis of our modelling results was tested for consistency with the crystal structures. In addition, the features stated to be essential for the basal activity based on the X-ray data were investigated by means of molecular dynamics simulations. Our results show that the homology modelling procedure was able to predict the CAR LBD structure with high accuracy. Structural features that have been revealed as critical for constitutive activity in the model are also observed in the X-ray structures. Furthermore, the MD simulations of the CAR X-ray structures and a detailed analysis of other NRs clarify the role of distinct structural features that have been assigned an important role for the constitutive activity.     

Homology modeling of the estrogen receptor subtype beta (ER-beta) and calculation of ligand binding affinities.

Estrogen is a steroid hormone playing critical roles in physiological processes such as sexual differentiation and development, female and male reproductive processes, and bone health. Numerous natural and synthetic environmental compounds have been shown capable of estrogenic effects. This area has been the focus of significant fundamental and applied research due both to the potential detrimental effects of these compounds upon normal physiological processes and to the potential beneficial effects of tissue-selective estrogen agonists/antagonists for the prevention and treatment of numerous diseases. Genomic effects of the active form of estrogen, 17beta-estradiol, are mediated through at least two members of the steroid hormone receptor superfamily, estrogen receptor subtype alpha (ER-alpha) and estrogen receptor subtype beta (ER-beta). At the time of this work, the X-ray crystal structure of the ER-alpha had been elucidated, however, coordinates of the ER-beta were not publicly available. Based upon the significant structural conservation across members of the steroid hormone receptor family, and the high sequence homology between ER-alpha and ER-beta (>60%), we have developed a homology model of the ER-beta structure. Using the crystal structure of ER-alpha and the homology model of ER-beta, we demonstrate a strong correlation between computed values of the binding-energy and published values of the observed relative binding affinity (RBA) for a variety of compounds for both receptors, as well as the ability to identify receptor subtype selective compounds. Furthermore, using the recently available crystal structure of ER-beta for comparison purposes, we show that not only is the predicted homology model structurally accurate, but that it can be used to assess ligand binding affinities.     

Homology modeling of the DNA-binding domain of human Smad5: a molecular model for inhibitor design

Members of the Smad protein family function as signal transducers of the transforming growth factor (TGF-beta) superfamily proteins. The human Smad5 protein, a signal transducer downstream of TGF-beta/BMP receptors, is composed of N-terminal DNA binding domain (MH1) and C-terminal protein-protein interaction domain (MH2) connected together by a linker motif. We used homology-modeling techniques to generate a reliable molecular model of the Smad5 MH1 domain based on the crystal structure of Smad3 MH1 domain. Our study presents the structural features of a BMP-regulated, R-Smad subfamily member (consisting of Smad1, Smad5 and Smad8) for the first time. This model provides a structural basis for explaining both functional similarities and differences between Smad3 and Smad5. Also, the structural model of this molecular target would be useful for structure-based inhibitor design because of its high accuracy. The results of our study provide important insights into understanding the structure-function relationship of the members of the Smad protein family and can serve to guide future genetic and biochemical experiments in this area.     

Structure and dynamics of 6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase

Folates are essential for life. Unlike mammals, most microorganisms must synthesize folates de novo. 6-Hydroxymethyl-7,8-dihydropterin pyrophosphokinase (HPPK) catalyzes pyrophosphoryl transfer from ATP to 6-hydroxymethyl-7,8-dihydropterin (HP), the first reaction in folate pathway, and therefore, is an ideal target for developing novel antimicrobial agents. Because of its small size and high thermal stability, E. coli HPPK is also an excellent model enzyme for studying the mechanisms of enzymatic pyrophosphoryl transfer. We have determined the crystal structures of HPPK in the unligated form and in complex with HP, two Mg2+ ions, and AMPCPP (an ATP analog that inhibits the enzymatic reaction). Comparison of the two crystal structures reveals dramatic conformational changes of three flexible loops and many side chains and possible roles of the active site residues.     

电子病历中基于XML的CDA应用

文章介绍了XML语言，它作为电子病历描述语言具有多项优点。它拥有大量的开发和应用工具，有利于对病历内容的处理。基于XML语言，医疗行业发布临床文档结构CDA，它提供了临床文件的交换模型，CDA为电子病历的实现提供了标准。文中通过实例说明基于XML的CDA的应用。     

2D clustering based discriminant analysis for 3D head model classification

This paper introduces a novel framework for 3D head model recognition based on the recently proposed 2D subspace analysis method. Two main contributions have been made. First, a 2D version of clustering-based discriminant analysis (CDA) is proposed, which combines the capability to model the multiple cluster structure embedded within a single class with the computational advantage that is characteristic of 2D subspace analysis methods. Second, we extend the applications of 2D subspace methods to the field of 3D head model classification by characterizing these models with 2D feature sets.     

An improved secondary structure computation method and its application to intervening sequence in the human alpha-like globin mRNA precursors

Current secondary structure prediction computations have a serious drawback. The calculated thermodynamically most stable structure often differs from that observed in solution or in crystal form. In this paper we suggest a way to partially over-come some of these limitations by simulating the RNA folding process and calculating the frequencies of occurrence of the various substructures obtained. The frequently recurring substructures are then selected to construct the secondary structure of the whole RNA. 142 tRNA molecules and an E. coli 16S rRNA molecule have been examined by this method. The percentage of successful prediction of the correct helices are significantly higher than those calculated previously. The secondary structures of intervening sequences (IVSs) excised from human alpha-like globin pre-mRNAs are also computed. Thus, in this method the secondary structures obtained are composed of the statistically more significant substructures. This has also been demonstrated by using randomly shuffled sequences. The secondary structures of each of the randomized sequences are computed and their mean and standard deviations are used in evaluating the significance of the substructures obtained in the folding of the biological sequence. Some potentially appealing structural features aligning adjacent exons for ligation have been found.     

Determining transmission line path alternatives using a valley-finding algorithm

Since new (Power) Transmission Lines (TLs) can have a long-term effect on the makeup of a landscape and on the human living space, it should be expected that the route of any new TL will be based on objective criteria that take into account the views of the public. Geographic Information Science (GIScience) provides powerful tools that assist in the determination of feasible locations for new TLs based on objective criteria and georeferenced data by combining methods from Multi-Criteria Decision Analysis (MCDA) and Least Cost Path (LCP) analysis. If such an approach is applied, the LCP analysis usually yields one optimal result. However, stakeholders and decision-makers prefer to compare multiple distinct path alternatives in order to find a solution that will be acceptable to as many stakeholders as possible.We have developed a method that calculates spatially distinct and Pareto optimal path alternatives based on the same cost surface using an algorithm for determining valleys on a Digital Elevation Model (DEM) to determine local low-cost points, which were then connected to a network graph by geometrical rules. Finally, we selected all non-dominated path alternatives that represented Pareto optimal conditions regarding a specific main objective. We then asked ten expert stakeholders to cross-compare the calculated path alternatives and assess our novel method. The concept of applying linear programming to obtain Pareto optimal path alternatives yielded routings that were mostly preferred over the LCP and had a greater likelihood of being realized than the results obtained by conducting the LCP analysis. The stakeholders determined the method's key concept to be useful and assert it a high potential to support planning, matter-of-fact argumentation, and discussions about TL routing.     

人手食指运动学建模

通过分析人手解剖学结构和观察人手运动特性，提出如下观点：（1）食指指掌关节具有两个轴线垂直相交的旋转自由度，两轴线相对于手掌不同的排列顺序对应于两种不同的运动学模型，指掌关节只能是其中的一种；（2）食指指掌关节的两个自由度在运动极限位置具有相关性．通过实验测绘出人手食指的工作空间形状，将之与两种可能的食指运动学模型对应的工作空间相比较，判别出指掌关节的真实运动学结构，并验证了指掌关节极限位置上两个自由度运动相关性．在上述工作基础上建立了符合人手实际结构的手指运动学模型．     

A scoring model to detect abusive billing patterns in health insurance claims

We propose a scoring model that detects outpatient clinics with abusive utilization patterns based on profiling information extracted from electronic insurance claims. The model consists of (1) scoring to quantify the degree of abusiveness and (2) segmentation to categorize the problematic providers with similar utilization patterns. We performed the modeling for 3705 Korean internal medicine clinics. We applied data from practitioner claims submitted to the National Health Insurance Corporation for outpatient care during the 3rd quarter of 2007 and used 4th quarter data to validate the model. We considered the Health Insurance Review and Assessment Services decisions on interventions to be accurate for model validation. We compared the conditional probability distributions of the composite degree of anomaly (CDA) score formulated for intervention and non-intervention groups. To assess the validity of the model, we examined confusion matrices by intervention history and group as defined by the CDA score. The CDA aggregated 38 indicators of abusiveness for individual clinics, which were grouped based on the CDAs, and we used the decision tree to further segment them into homogeneous clusters based on their utilization patterns. The validation indicated that the proposed model was largely consistent with the manual detection techniques currently used to identify potential abusers. The proposed model, which can be used to automate abuse detection, is flexible and easy to update. It may present an opportunity to fight escalating healthcare costs in the era of increasing availability of electronic healthcare information.     

A combined QM and MM investigation into guanine quadruplexes

This paper reports on the application of quantum mechanical (QM) energy calculations, QM optimisations and MD simulations to explore the stability of a human telomeric guanine quadruplex, containing potassium and sodium cations. G-quadruplexes are of great biological interest as it has been suggested that they offer a novel path to cancer inhibition. By understanding the stability and geometry of these DNA features gives us the ability to design ligands which can bind and stabilise the G-quadruplex. There are significant structural differences between the potassium containing crystal structure of human telomeric G-quadruplex and the sodium containing NMR structure; in this paper, we investigate the energetics and dynamics of the potassium derived crystal structure and a model for the sodium containing structure. QM investigations upon the 12 G-quadruplex core, extracted from the human potassium quadruplex crystal structure, indicate that replacement of the potassium cations with sodium yields an energetically more favourable structure. However, attempts to geometry optimise both structures at the QM level proved unsuccessful, the structure of the partially optimised potassium containing G-quadruplex retains significant structural integrity with respect to the original crystal structure, whilst the sodium containing G-quadruplex shows significant structural distortion. QM investigation of the 12 G-quadruplex core containing no cations unsurprisingly yields a highly unfavourable energetic structure. MD simulations on the complete quadruplex structure, containing potassium cations, yields a remarkably stable structure after 4ns of simulation, the most significant deviation from the original crystal structure being the loss of the capping potassium cation from the structure. MD simulation of the sodium containing quadruplex for 4ns show significant structural reorganisation compared with the original potassium containing crystal structure.     

电子病历中基于XML的CDA应用

文章介绍了XML语言,它作为电子病历描述语言具有多项优点。它拥有大量的开发和应用工具,有利于对病历内容的处理。基于XML语言,医疗行业发布临床文档结构CDA,它提供了临床文件的交换模型,CDA为电子病历的实现提供了标准。文中通过实例说明基于XML的CDA的应用。     

A proposal for modelling usability in a holistic MDD method

Holistic methods for Model-Driven Development (MDD) aim to model all the system features in a conceptual model. This conceptual model is the input for a model compiler that can generate software systems by means of automatic transformations. However, in general, MDD methods focus on modelling the structure and functionality of systems, relegating the interaction and usability features to manual implementations at the last steps of the software development process. Some usability features are strongly related to the functionality of the system and their inclusion is not so easy. In order to facilitate the inclusion of functional usability features from the first steps of the development process and bring closer MDD methods to the holistic perspective, we propose a Usability Model. The Usability Model gathers conceptual primitives that represent functional usability features in a sufficiently abstract way so that the model can be used with different holistic MDD methods. This paper defines all the primitives that can be used to represent functional usability features. Moreover, we have defined a process to include the Usability Model in any MDD method without affecting its existing conceptual model. The proposal is based on model-to-model and model-to-code transformations. As proof of concept, we have applied our proposal to an existing MDD method called the OO-method and we have measured its efficiency.     

Molecular basis of the constitutive activity and STI571 resistance of Asp816Val mutant KIT receptor tyrosine kinase

The receptor tyrosine kinase, KIT, displays activating mutations in the kinase domain, which are associated with various cancers. We have used homology modelling based on the crystal structures of the insulin receptor kinase in active and inactive conformations to predict the corresponding structures of the KIT kinase domain. We have prepared four KIT models, one each for the active and inactive conformations of the wild-type and of the Asp816Val mutant proteins. We have also placed ATP into the active conformations and the inhibitor, STI571, into the inactive conformations. All models have been fully energy minimised. The molecular modelling studies described here explain (i) why Asp816Val KIT is constitutively active, (ii) why the nature of the substituting amino acid at residue 816 is relatively unimportant, and (iii) why the Asp816Val substitution confers resistance to the KIT-inhibitory drug STI571. The models will be valuable for predicting other kinase inhibitory drugs that may be active on wild-type and mutant forms of KIT. During the course of this work, a crystal structure of the active conformation of the KIT kinase domain has been published. Our model of the active conformation of the Asp816Val mutant is strikingly similar to this crystal structure, whereas our model of the active conformation of the wild-type kinase domain of KIT differs from the crystal structure in some respects. The reasons for this apparent discrepancy are discussed.