Search PubMed⌕ Search

Biomedical subjects

Andrej Sali

Publications and source records attributed to Andrej Sali.

At least 19 recordsLinked to original sources
PubMed2006

Structural modeling of protein interactions by analogy: application to PSD-95.

We describe comparative patch analysis for modeling the structures of multidomain proteins and protein complexes, and apply it to the PSD-95 protein. Comparative patch analysis is a hybrid of comparative modeling based on a template complex and protein docking, with a greater applicability than comparative modeling and a higher accuracy than docking. It relies on structurally defined interactions of each of the complex components, or their homologs, with any other protein, irrespective of its fold. For each component, its known binding modes with other proteins of any fold are collected and expanded by the known binding modes of its homologs. These modes are then used to restrain conventional molecular docking, resulting in a set of binary domain complexes that are subsequently ranked by geometric complementarity and a statistical potential. The method is evaluated by predicting 20 binary complexes of known structure. It is able to correctly identify the binding mode in 70% of the benchmark complexes compared with 30% for protein docking. We applied comparative patch analysis to model the complex of the third PSD-95, DLG, and ZO-1 (PDZ) domain and the SH3-GK domains in the PSD-95 protein, whose structure is unknown. In the first predicted configuration of the domains, PDZ interacts with SH3, leaving both the GMP-binding site of guanylate kinase (GK) and the C-terminus binding cleft of PDZ accessible, while in the second configuration PDZ interacts with GK, burying both binding sites. We suggest that the two alternate configurations correspond to the different functional forms of PSD-95 and provide a possible structural description for the experimentally observed cooperative folding transitions in PSD-95 and its homologs. More generally, we expect that comparative patch analysis will provide useful spatial restraints for the structural characterization of an increasing number of binary and higher-order protein complexes.

Amino Acid Sequence↗
PubMed2006

Localization of binding sites in protein structures by optimization of a composite scoring function.

The rise in the number of functionally uncharacterized protein structures is increasing the demand for structure-based methods for functional annotation. Here, we describe a method for predicting the location of a binding site of a given type on a target protein structure. The method begins by constructing a scoring function, followed by a Monte Carlo optimization, to find a good scoring patch on the protein surface. The scoring function is a weighted linear combination of the z-scores of various properties of protein structure and sequence, including amino acid residue conservation, compactness, protrusion, convexity, rigidity, hydrophobicity, and charge density; the weights are calculated from a set of previously identified instances of the binding-site type on known protein structures. The scoring function can easily incorporate different types of information useful in localization, thus increasing the applicability and accuracy of the approach. To test the method, 1008 known protein structures were split into 20 different groups according to the type of the bound ligand. For nonsugar ligands, such as various nucleotides, binding sites were correctly identified in 55%-73% of the cases. The method is completely automated (http://salilab.org/patcher) and can be applied on a large scale in a structural genomics setting.

Amino Acid Sequence↗
PubMed2006

Minimalist representations and the importance of nearest neighbor effects in protein folding simulations.

In order to investigate the level of representation required to simulate folding and predict structure, we test the ability of a variety of reduced representations to identify native states in decoy libraries and to recover the native structure given the advanced knowledge of the very broad native Ramachandran basin assignments. Simplifications include the removal of the entire side-chain or the retention of only the Cbeta atoms. Scoring functions are derived from an all-atom statistical potential that distinguishes between atoms and different residue types. Structures are obtained by minimizing the scoring function with a computationally rapid simulated annealing algorithm. Results are compared for simulations in which backbone conformations are sampled from a Protein Data Bank-based backbone rotamer library generated by either ignoring or including a dependence on the identity and conformation of the neighboring residues. Only when the Cbeta atoms and nearest neighbor effects are included do the lowest energy structures generally fall within 4 A of the native backbone root-mean square deviation (RMSD), despite the initial configuration being highly expanded with an average RMSD > or = 10 A. The side-chains are reinserted into the Cbeta models with minimal steric clash. Therefore, the detailed, all-atom information lost in descending to a Cbeta-level representation is recaptured to a large measure using backbone dihedral angle sampling that includes nearest neighbor effects and an appropriate scoring function.

Amino Acids↗
PubMed2006

A composite score for predicting errors in protein structure models.

Reliable prediction of model accuracy is an important unsolved problem in protein structure modeling. To address this problem, we studied 24 individual assessment scores, including physics-based energy functions, statistical potentials, and machine learning-based scoring functions. Individual scores were also used to construct approximately 85,000 composite scoring functions using support vector machine (SVM) regression. The scores were tested for their abilities to identify the most native-like models from a set of 6000 comparative models of 20 representative protein structures. Each of the 20 targets was modeled using a template of <30% sequence identity, corresponding to challenging comparative modeling cases. The best SVM score outperformed all individual scores by decreasing the average RMSD difference between the model identified as the best of the set and the model with the lowest RMSD (DeltaRMSD) from 0.63 A to 0.45 A, while having a higher Pearson correlation coefficient to RMSD (r=0.87) than any other tested score. The most accurate score is based on a combination of the DOPE non-hydrogen atom statistical potential; surface, contact, and combined statistical potentials from MODPIPE; and two PSIPRED/DSSP scores. It was implemented in the SVMod program, which can now be applied to select the final model in various modeling problems, including fold assignment, target-template alignment, and loop modeling.

Models, Molecular↗
PubMed2006

Protein complex compositions predicted by structural similarity.

Proteins function through interactions with other molecules. Thus, the network of physical interactions among proteins is of great interest to both experimental and computational biologists. Here we present structure-based predictions of 3387 binary and 1234 higher order protein complexes in Saccharomyces cerevisiae involving 924 and 195 proteins, respectively. To generate candidate complexes, comparative models of individual proteins were built and combined together using complexes of known structure as templates. These candidate complexes were then assessed using a statistical potential, derived from binary domain interfaces in PIBASE (http://salilab.org/pibase). The statistical potential discriminated a benchmark set of 100 interface structures from a set of sequence-randomized negative examples with a false positive rate of 3% and a true positive rate of 97%. Moreover, the predicted complexes were also filtered using functional annotation and sub-cellular localization data. The ability of the method to select the correct binding mode among alternates is demonstrated for three camelid VHH domain-porcine alpha-amylase interactions. We also highlight the prediction of co-complexed domain superfamilies that are not present in template complexes. Through integration with MODBASE, the application of the method to proteomes that are less well characterized than that of S.cerevisiae will contribute to expansion of the structural and functional coverage of protein interaction space. The predicted complexes are deposited in MODBASE (http://salilab.org/modbase).

Algorithms↗
PubMed2006

Simple fold composition and modular architecture of the nuclear pore complex.

The nuclear pore complex (NPC) consists of multiple copies of approximately 30 different proteins [nucleoporins (nups)], forming a channel in the nuclear envelope that mediates macromolecular transport between the cytosol and the nucleus. With <5% of the nup residues currently available in experimentally determined structures, little is known about the detailed structure of the NPC. Here, we use a combined computational and biochemical approach to assign folds for approximately 95% of the residues in the yeast and vertebrate nups. These fold assignments suggest an underlying simplicity in the composition and modularity in the architecture of all eukaryotic NPCs. The simplicity in NPC composition is reflected in the presence of only eight fold types, with the three most frequent folds accounting for approximately 85% of the residues. The modularity in NPC architecture is reflected in its hierarchical and symmetrical organization that partitions the predicted nup folds into three groups: the transmembrane group containing transmembrane helices and a cadherin fold, the central scaffold group containing beta-propeller and alpha-solenoid folds, and the peripheral FG group containing predominantly the FG repeats and the coiled-coil fold. Moreover, similarities between structures in coated vesicles and those in the NPC support our prior hypothesis for their common evolutionary origin in a progenitor protocoatomer. The small number of predicted fold types in the NPC and their internal symmetries suggest that the bulk of the NPC structure has evolved through extensive motif and gene duplication from a simple precursor set of only a few proteins.

Computational Biology↗
PubMed2006

Refinement of protein structures by iterative comparative modeling and CryoEM density fitting.

We developed a method for structure characterization of assembly components by iterative comparative protein structure modeling and fitting into cryo-electron microscopy (cryoEM) density maps. Specifically, we calculate a comparative model of a given component by considering many alternative alignments between the target sequence and a related template structure while optimizing the fit of a model into the corresponding density map. The method relies on the previously developed Moulder protocol that iterates over alignment, model building, and model assessment. The protocol was benchmarked using 20 varied target-template pairs of known structures with less than 30% sequence identity and corresponding simulated density maps at resolutions from 5A to 25A. Relative to the models based on the best existing sequence profile alignment methods, the percentage of C(alpha) atoms that are within 5A of the corresponding C(alpha) atoms in the superposed native structure increases on average from 52% to 66%, which is half-way between the starting models and the models from the best possible alignments (82%). The test also reveals that despite the improvements in the accuracy of the fitness function, this function is still the bottleneck in reducing the remaining errors. To demonstrate the usefulness of the protocol, we applied it to the upper domain of the P8 capsid protein of rice dwarf virus that has been studied by cryoEM at 6.8A. The C(alpha) root-mean-square deviation of the model based on the remotely related template, bluetongue virus VP7, improved from 8.7A to 6.0A, while the best possible model has a C(alpha) RMSD value of 5.3A. Moreover, the resulting model fits better into the cryoEM density map than the initial template structure. The method is being implemented in our program MODELLER for protein structure modeling by satisfaction of spatial restraints and will be applicable to the rapidly increasing number of cryoEM density maps of macromolecular assemblies.

Amino Acid Sequence↗
PubMed2006

Variable gap penalty for protein sequence-structure alignment.

The penalty for inserting gaps into an alignment between two protein sequences is a major determinant of the alignment accuracy. Here, we present an algorithm for finding a globally optimal alignment by dynamic programming that can use a variable gap penalty (VGP) function of any form. We also describe a specific function that depends on the structural context of an insertion or deletion. It penalizes gaps that are introduced within regions of regular secondary structure, buried regions, straight segments and also between two spatially distant residues. The parameters of the penalty function were optimized on a set of 240 sequence pairs of known structure, spanning the sequence identity range of 20-40%. We then tested the algorithm on another set of 238 sequence pairs of known structures. The use of the VGP function increases the number of correctly aligned residues from 81.0 to 84.5% in comparison with the optimized affine gap penalty function; this difference is statistically significant according to Student's t-test. We estimate that the new algorithm allows us to produce comparative models with an additional approximately 7 million accurately modeled residues in the approximately 1.1 million proteins that are detectably related to a known structure.

Algorithms↗
PubMed2006

MODBASE: a database of annotated comparative protein structure models and associated resources.

MODBASE (http://salilab.org/modbase) is a database of annotated comparative protein structure models for all available protein sequences that can be matched to at least one known protein structure. The models are calculated by MODPIPE, an automated modeling pipeline that relies on MODELLER for fold assignment, sequence-structure alignment, model building and model assessment (http:/salilab.org/modeller). MODBASE is updated regularly to reflect the growth in protein sequence and structure databases, and improvements in the software for calculating the models. MODBASE currently contains 3 094 524 reliable models for domains in 1 094 750 out of 1 817 889 unique protein sequences in the UniProt database (July 5, 2005); only models based on statistically significant alignments and models assessed to have the correct fold despite insignificant alignments are included. MODBASE also allows users to generate comparative models for proteins of interest with the automated modeling server MODWEB (http://salilab.org/modweb). Our other resources integrated with MODBASE include comprehensive databases of multiple protein structure alignments (DBAli, http://salilab.org/dbali), structurally defined ligand binding sites and structurally defined binary domain interfaces (PIBASE, http://salilab.org/pibase) as well as predictions of ligand binding sites, interactions between yeast proteins, and functional consequences of human nsSNPs (LS-SNP, http://salilab.org/LS-SNP).

Binding Sites↗
PubMed2006

Protease accessibility laddering: a proteomic tool for probing protein structure.

Limited proteolysis is widely used in biochemical and crystallographic studies to determine domain organization, folding properties, and ligand binding activities of proteins. The method has limitations, however, due to the difficulties in obtaining sufficient amounts of correctly folded proteins and in interpreting the results of the proteolysis. A new limited proteolysis method, named protease accessibility laddering (PAL), avoids these complications. In PAL, tagged proteins are purified on magnetic beads in their natively folded state. While attached to the beads, proteins are probed with proteases. Proteolytic fragments are eluted and detected by immunoblotting with antibodies against the tag (e.g., Protein A, GFP, and 6xHis). PAL readily detects domain boundaries and flexible loops within proteins. A combination of PAL and comparative protein structure modeling allows characterization of previously unknown structures (e.g., Sec31, a component of the COPII coated vesicle). PAL's high throughput should greatly facilitate structural genomic and proteomic studies.

Adaptor Proteins, Signal Transducing↗
PubMed2006

Statistical potential for assessment and prediction of protein structures.

Protein structures in the Protein Data Bank provide a wealth of data about the interactions that determine the native states of proteins. Using the probability theory, we derive an atomic distance-dependent statistical potential from a sample of native structures that does not depend on any adjustable parameters (Discrete Optimized Protein Energy, or DOPE). DOPE is based on an improved reference state that corresponds to noninteracting atoms in a homogeneous sphere with the radius dependent on a sample native structure; it thus accounts for the finite and spherical shape of the native structures. The DOPE potential was extracted from a nonredundant set of 1472 crystallographic structures. We tested DOPE and five other scoring functions by the detection of the native state among six multiple target decoy sets, the correlation between the score and model error, and the identification of the most accurate non-native structure in the decoy set. For all decoy sets, DOPE is the best performing function in terms of all criteria, except for a tie in one criterion for one decoy set. To facilitate its use in various applications, such as model assessment, loop modeling, and fitting into cryo-electron microscopy mass density maps combined with comparative protein structure modeling, DOPE was incorporated into the modeling package MODELLER-8.

Computational Biology↗
PubMed2005

The C-type lectin fold as an evolutionary solution for massive sequence variation.

Only few instances are known of protein folds that tolerate massive sequence variation for the sake of binding diversity. The most extensively characterized is the immunoglobulin fold. We now add to this the C-type lectin (CLec) fold, as found in the major tropism determinant (Mtd), a retroelement-encoded receptor-binding protein of Bordetella bacteriophage. Variation in Mtd, with its approximately 10(13) possible sequences, enables phage adaptation to Bordetella spp. Mtd is an intertwined, pyramid-shaped trimer, with variable residues organized by its CLec fold into discrete receptor-binding sites. The CLec fold provides a highly static scaffold for combinatorial display of variable residues, probably reflecting a different evolutionary solution for balancing diversity against stability from that in the immunoglobulin fold. Mtd variants are biased toward the receptor pertactin, and there is evidence that the CLec fold is used broadly for sequence variation by related retroelements.

Amino Acid Sequence↗
PubMed2005

Localization of protein-binding sites within families of proteins.

We address the question of whether or not the positions of protein-binding sites on homologous protein structures are conserved irrespective of the identities of their binding partners. First, for each domain family in the Structural Classification of Proteins (SCOP), protein-binding sites are extracted from our comprehensive database of structurally defined binary domain interactions (PIBASE). Second, the binding sites within each family are superposed using a structural alignment of its members. Finally, the degree of localization of binding sites within each family is quantified by comparing it with localization expected by chance. We found that 72% of the 1847 SCOP domain families in PIBASE have binding sites with localization values greater than expected by chance. Moreover, 554 (30%) of these families have localizations that are statistically significant (i.e., more than four standard deviations away from the mean expected by chance). In contrast, only 144 (8%) families have significantly low localization. The absence of a significant correlation of the binding site localization with the average sequence and structural conservations in a family suggests that localization can be helpful for describing the functional diversity of protein-protein interactions, complementing measures of sequence and structural conservation. Consideration of the binding site localization may also result in spatial restraints for the modeling of protein assembly structures.

Acyl-CoA Dehydrogenase↗
PubMed2005

Detecting remotely related proteins by their interactions and sequence similarity.

The function of an uncharacterized protein is usually inferred either from its homology to, or its interactions with, characterized proteins. Here, we use both sequence similarity and protein interactions to identify relationships between remotely related protein sequences. We rely on the fact that homologous sequences share similar interactions, and, therefore, the set of interacting partners of the partners of a given protein is enriched by its homologs. The approach was bench-marked by assigning the fold and functional family to test sequences of known structure. Specifically, we relied on 1,434 proteins with known folds, as defined in the Structural Classification of Proteins (SCOP) database, and with known interacting partners, as defined in the Database of Interacting Proteins (DIP). For this subset, the specificity of fold assignment was increased from 54% for position-specific iterative BLAST to 75% for our approach, with a concomitant increase in sensitivity for a few percentage points. Similarly, the specificity of family assignment at the e-value threshold of 10(-8) was increased from 70% to 87%. The proposed method would be a useful tool for large-scale automated discovery of remote relationships between protein sequences, given its unique reliance on sequence similarity and protein-protein interactions.

Computational Biology↗
PubMed2005

LS-SNP: large-scale annotation of coding non-synonymous SNPs based on multiple information sources.

MOTIVATION: The NCBI dbSNP database lists over 9 million single nucleotide polymorphisms (SNPs) in the human genome, but currently contains limited annotation information. SNPs that result in amino acid residue changes (nsSNPs) are of critical importance in variation between individuals, including disease and drug sensitivity. RESULTS: We have developed LS-SNP, a genomic scale software pipeline to annotate nsSNPs. LS-SNP comprehensively maps nsSNPs onto protein sequences, functional pathways and comparative protein structure models, and predicts positions where nsSNPs destabilize proteins, interfere with the formation of domain-domain interfaces, have an effect on protein-ligand binding or severely impact human health. It currently annotates 28,043 validated SNPs that produce amino acid residue substitutions in human proteins from the SwissProt/TrEMBL database. Annotations can be viewed via a web interface either in the context of a genomic region or by selecting sets of SNPs, genes, proteins or pathways. These results are useful for identifying candidate functional SNPs within a gene, haplotype or pathway and in probing molecular mechanisms responsible for functional impacts of nsSNPs. AVAILABILITY: http://www.salilab.org/LS-SNP CONTACT: rachelk@salilab.org SUPPLEMENTARY INFORMATION: http://salilab.org/LS-SNP/supp-info.pdf.

Algorithms↗
PubMed2005

Urokinase-type plasminogen activator is a preferred substrate of the human epithelium serine protease tryptase epsilon/PRSS22.

Tryptase epsilon is a member of the chromosome 16p13.3 family of human serine proteases that is preferentially expressed by epithelial cells. Recombinant pro-tryptase epsilon was generated to understand how the exocytosed zymogen might be activated outside of the epithelial cell, as well as to address its possible role in normal and diseased states. Using expression/site-directed mutagenesis approaches, we now show that Lys20, Cys90, and Asp92 in the protease's substrate-binding cleft regulate its enzymatic activity. We also show that Arg(-1) in the propeptide domain controls its ability to autoactivate. In vitro studies revealed that recombinant tryptase epsilon possesses a restricted substrate specificity. Once activated, tryptase epsilon cannot be inhibited effectively by the diverse array of protease inhibitors present in normal human plasma. Moreover, this epithelium protease is not highly susceptible to alpha1-antitrypsin or secretory leukocyte protease inhibitor, which are present in the lung. Recombinant tryptase epsilon could not cleave fibronectin, vitronectin, laminin, single-chain tissue-type plasminogen activator, plasminogen, or any prominent serum protein. Nevertheless, tryptase epsilon readily converted single-chain pro-urokinase-type plasminogen activator (pro-uPA/scuPA) into its mature, enzymatically active protease. Tryptase epsilon also was able to induce pro-uPA-expressing smooth muscle cells to increase their migration through a basement membrane-like extracellular matrix. The ability to activate uPA in the presence of varied protease inhibitors suggests that tryptase epsilon plays a prominent role in fibrinolysis and other uPA-dependent reactions in the lung.

Animals↗