Joint Center For Structural Genomics Research Articles

Buccaneer model building with neural network fragment selection.

Tracing the backbone is a critical step in protein model building, as incorrect tracing leads to poor protein models. Here, a neural network trained to identify unfavourable fragments and remove them from the model-building process in order to improve backbone tracing is presented. Moreover, a decision tree was trained to select an optimal threshold to eliminate unfavourable fragments. The neural network was tested on experimental phasing data sets from the Joint Center for Structural Genomics (JCSG), recently deposited experimental phasing data sets (from 2015 to 2021) and molecular-replacement data sets. The experimental results show that using the neural network in the Buccaneer protein-model-building software can produce significantly more complete protein models than those built using Buccaneer alone. In particular, Buccaneer with the neural network built protein models with a completeness that was at least 5% higher for 25% and 50% of the original and truncated resolution JCSG experimental phasing data sets, respectively, for 28% of the recently collected experimental phasing data sets and for 43% of the molecular-replacement data sets.

NMR in structural genomics to increase structural coverage of the protein universe: Delivered by Prof. Kurt Wüthrich on 7 July 2013 at the 38th FEBS Congress in St. Petersburg, Russia.

For more than a decade, the Joint Center for Structural Genomics (JCSG; www.jcsg.org) worked toward increased three-dimensional structure coverage of the protein universe. This coordinated quest was one of the main goals of the four high-throughput (HT) structure determination centers of the Protein Structure Initiative (PSI; www.nigms.nih.gov/Research/specificareas/PSI). To achieve the goals of the PSI, the JCSG made use of the complementarity of structure determination by X-ray crystallography and nuclear magnetic resonance (NMR) spectroscopy to increase and diversify the range of targets entering the HT structure determination pipeline. The overall strategy, for both techniques, was to determine atomic resolution structures for representatives of large protein families, as defined by the Pfam database, which had no structural coverage and could make significant contributions to biological and biomedical research. Furthermore, the experimental structures could be leveraged by homology modeling to further expand the structural coverage of the protein universe and increase biological insights. Here, we describe what could be achieved by this structural genomics approach, using as an illustration the contributions from 20 NMR structure determinations out of a total of 98 JCSG NMR structures, which were selected because they are the first three-dimensional structure representations of the respective Pfam protein families. The information from this small sample is representative for the overall results from crystal and NMR structure determination in the JCSG. There are five new folds, which were classified as domains of unknown functions (DUF), three of the proteins could be functionally annotated based on three-dimensional structure similarity with previously characterized proteins, and 12 proteins showed only limited similarity with previous deposits in the Protein Data Bank (PDB) and were classified as DUFs.

Macromolecular crystallography and what it can contribute to antiparasite drug discovery.

This issue of Acta Crystallographica Section F is dedicated towards structural investigations on proteins associated with molecular parasitology, specifically research linked to protozoan pathogens. It is in line with the previous special sections or issues contributed by the RIKEN-UK structural genomics consortium (December 2009), the Joint Center for Structural Genomics in October 2010 (Weiss & Einspahr, 2010 ▶) and the Seattle Structural Genomics Center for Infectious Diseases in September 2011 (Weiss & Einspahr, 2011 ▶). In contrast to the previous issues however, it is thematically much more focused on protozoan pathogens. Interest in these primitive eukaryotic organisms is often to exploit them as model systems and many significant advances have resulted from such research. As examples there are the fundamentally important discoveries relating to RNA editing, antigenic variation and the occurrence of glycosylphosphatidylinositol tags that anchor proteins on the cell surface, all accrued from research with the African trypanosome. However, a major impetus to study protists is the drastic toll they take on human life. The World Health Organization provide documentation and sobering statistics on the diseases and their impact on human health. For example http://www.who.int/mediacentre/factsheets/fs375/en/ is their recently updated information sheet on Leishmaniasis. Researchers have sought to understand the processes that allow pathogens to exist, to invade a host, to evade the immune response, and to cause debilitating and often fatal diseases. Extending from such investigations and alongside drug discovery efforts enter structural biology, in particular crystallography and the high quality chemical information it can provide. Remember that most drugs are smallish organic molecules that exert their beneficial effect by interacting with certain macromolecules. The discovery of such molecules has transformed human existence. Indeed the emergence of the modern pharmaceutical industry, founded as it was on the basis of antimicrobial and analgesic agents, might be considered one of the major developments of the last 100 years. Different disciplines contribute to drug discovery, and increasingly the search is on for new targets, better technologies and model systems to assist the process. Structural studies contribute a key role by providing accurate chemical information to guide assessment of targets, the development of reliable structure–activity relationships or templates for computational design projects in the search for lead compounds. There are numerous excellent studies published in which specific parasite systems have been targeted and dissected. In addition structural genomics efforts have involved cohorts of protist proteins and the Protein Data Bank has been greatly enriched in this area with both NMR and crystal structures. We now present an issue of fundamental studies in molecular parasitology and also research directed toward protozoan drug target characterization. To set the scene there is an overview and perspective from Professor Wim Hol. In this he also details a few of his favourite examples where structural information contributes to the search for improved therapies. We commissioned Wim for an overview because of his outstanding contributions and infectious enthusiasm for the area; he continues to be an inspirational champion for the role of crystallography in drug discovery. There is also a short review from Professor Inari Kursula on cytoskeletal proteins related to the infection process. We thought it relevant to address this aspect and would highlight this as one in which there may be opportunities in future drug discovery work. We are pleased to also have a number of communications from structural genomics efforts which sit comfortably alongside more focused projects. Section F’s mandate is to allow the structural biology community to efficiently publish their data and we hope this special issue will serve to inform our community and perhaps others in the life sciences or medicinal chemistry areas, generate interest and provide encouragement to consider the journal as a natural home for results in molecular parasitology. We look forward to seeing more structural information on parasite proteins, more advances in basic biology and results to support future efforts in early stage drug discovery.

APSY-NMR for protein backbone assignment in high-throughput structural biology.

A standard set of three APSY-NMR experiments has been used in daily practice to obtain polypeptide backbone NMR assignments in globular proteins with sizes up to about 150 residues, which had been identified as targets for structure determination by the Joint Center for Structural Genomics (JCSG) under the auspices of the Protein Structure Initiative (PSI). In a representative sample of 30 proteins, initial fully automated data analysis with the software UNIO-MATCH-2014 yielded complete or partial assignments for over 90 % of the residues. For most proteins the APSY data acquisition was completed in less than 30 h. The results of the automated procedure provided a basis for efficient interactive validation and extension to near-completion of the assignments by reference to the same 3D heteronuclear-resolved [(1)H,(1)H]-NOESY spectra that were subsequently used for the collection of conformational constraints. High-quality structures were obtained for all 30 proteins, using the J-UNIO protocol, which includes extensive automation of NMR structure determination.

JCSG – Adapting Structural Genomics to Eukaryotic Complexes

For over a decade, the Joint Center for Structural Genomics (JCSG.org) has been at the forefront of developing tools and methodologies that enable the application of high-throughput structural biology (HTBSB) approaches to a broad range of challenging biological and biomedical problems. In PSI:Biology (2010-2015) to meet the challenges and embrace the opportunities that arise from our Partnerships projects, we have leveraged our gene-to-structure pipeline to explore challenging projects focused on structural characterization of interaction networks involved in stem cell regulation, T-cell activation and nuclear receptor signaling. These highly collaborative efforts have enabled development of systematic and integrative approaches for identifying and investigating networks of key multi-domain eukaryotic proteins and higher order assemblies of multi-component eukaryotic protein/protein and protein/nucleic acid complexes. In parallel, our biomedical theme project has focused on investigating host/microbe interactions of the microbial communities that inhabit specific niches and environments of the human body, e.g. the human gut microbiome. These efforts to date have been centered on secreted proteins from commensal bacteria in the human gut. The symbiotic relationship and influence on human development, physiology, immunity, and nutrition represent an exciting new frontier for HTBSB where we can investigate how these microorganisms contribute to human health as well as to disease. The JCSG also strives to promote widespread use of PSI resources, materials, methodologies and data to the general scientific community, via Community Nominated Target (CNT) projects and development and use of new technologies and methodologies. We also continue to contribute to the original PSI mission of structural coverage of the expanding protein universe. Supported by NIH U54-GM094586.

Improving the chances of successful protein structure determination with a random forest classifier.

Obtaining diffraction quality crystals remains one of the major bottlenecks in structural biology. The ability to predict the chances of crystallization from the amino-acid sequence of the protein can, at least partly, address this problem by allowing a crystallographer to select homologs that are more likely to succeed and/or to modify the sequence of the target to avoid features that are detrimental to successful crystallization. In 2007, the now widely used XtalPred algorithm [Slabinski et al. (2007), Protein Sci. 16, 2472-2482] was developed. XtalPred classifies proteins into five `crystallization classes' based on a simple statistical analysis of the physicochemical features of a protein. Here, towards the same goal, advanced machine-learning methods are applied and, in addition, the predictive potential of additional protein features such as predicted surface ruggedness, hydrophobicity, side-chain entropy of surface residues and amino-acid composition of the predicted protein surface are tested. The new XtalPred-RF (random forest) achieves significant improvement of the prediction of crystallization success over the original XtalPred. To illustrate this, XtalPred-RF was tested by revisiting target selection from 271 Pfam families targeted by the Joint Center for Structural Genomics (JCSG) in PSI-2, and it was estimated that the number of targets entered into the protein-production and crystallization pipeline could have been reduced by 30% without lowering the number of families for which the first structures were solved. The prediction improvement depends on the subset of targets used as a testing set and reaches 100% (i.e. twofold) for the top class of predicted targets.

New mini- zincin structures provide a minimal scaffold for members of this metallopeptidase superfamily

BackgroundThe Acel_2062 protein from Acidothermus cellulolyticus is a protein of unknown function. Initial sequence analysis predicted that it was a metallopeptidase from the presence of a motif conserved amongst the Asp-zincins, which are peptidases that contain a single, catalytic zinc ion ligated by the histidines and aspartic acid within the motif (HEXXHXXGXXD). The Acel_2062 protein was chosen by the Joint Center for Structural Genomics for crystal structure determination to explore novel protein sequence space and structure-based function annotation.ResultsThe crystal structure confirmed that the Acel_2062 protein consisted of a single, zincin-like metallopeptidase-like domain. The Met-turn, a structural feature thought to be important for a Met-zincin because it stabilizes the active site, is absent, and its stabilizing role may have been conferred to the C-terminal Tyr113. In our crystallographic model there are two molecules in the asymmetric unit and from size-exclusion chromatography, the protein dimerizes in solution. A water molecule is present in the putative zinc-binding site in one monomer, which is replaced by one of two observed conformations of His95 in the other.ConclusionsThe Acel_2062 protein is structurally related to the zincins. It contains the minimum structural features of a member of this protein superfamily, and can be described as a “mini- zincin”. There is a striking parallel with the structure of a mini-Glu-zincin, which represents the minimum structure of a Glu-zincin (a metallopeptidase in which the third zinc ligand is a glutamic acid). Rather than being an ancestral state, phylogenetic analysis suggests that the mini-zincins are derived from larger proteins.

The Structure of an Aldehyde Decarbonylase from Prochlorococcus Marinus Implicated in Biofuel Synthesis Indicates an Octadecanoic Acid Ligand Coordinated to a Di-Iron Catalytic Center∗

Prochlorococcus marinus, a prevalent marine cyanobacterium, can use sunlight and atmospheric CO2 to convert fatty-acid like precursors into alkane biofuels. Schirmer et al. (Science, 2010) showed that alkane production in cyanobacteria entails a two-enzyme process in which the acyl-CoA moiety is reduced to an aldehyde followed by decarbonylation to an alkane. The structure of the aldehyde decarbonylase has been determined by the Joint Center for Structural Genomics (PDB ID 2oc5). Here, we have identified the bound metals at the catalytic site as Fe and, based on the results of crystallographic refinement, have assigned a previously unidentified ligand as an octadecanoic acid. The metal identity was determined from X-ray fluorescence and anomalous difference Fourier maps. The oxygen atoms of the octadecanoic acid are within coordination distance of the Fe atoms and the hydrocarbon tail extends into the interior of the protein. Previous studies showed the enzyme can use octadecanal as a substrate, but the structure raises the question if it can directly catalyze alkane formation from octadecanoic acid ∗Supported by NIGMS PSI grants U54 GM094586 and GM074898View Large Image | View Hi-Res Image | Download PowerPoint Slide

The Joint Center for Structural Genomics

The Joint Center for Structural Genomics

PSI-Biology partnership projects at the Joint Center for Structural Genomics

PSI-Biology partnership projects at the Joint Center for Structural Genomics

Cases of twinning at the Joint Center for Structural Genomics

Cases of twinning at the Joint Center for Structural Genomics

Community nominated projects at the Joint Center for Structural Genomics

Community nominated projects at the Joint Center for Structural Genomics

NMR structure of the Bordetella bronchiseptica protein NP_888769.1 establishes a new phage-related protein family PF13554

The solution structure of the hypothetical phage-related protein NP_888769.1 from the Gram-negative bacterium Bordetella bronchoseptica contains a well-structured core comprising a five-stranded, antiparallel β-sheet packed on one side against two α-helices and a short β-hairpin with three flexibly disordered loops extending from the central β-sheet. A homology search with the software DALI identified two Protein Data Bank deposits with Z-scores > 8, where both of these proteins have less than 8% sequence identity relative to NP_888769.1, and one has been functionally annotated as a lambda phage tail terminator protein. A sequence-homology analysis then confirmed that NP_888769.1 represents the first three-dimensional structural representative of a new protein family that was previously predicted by the Joint Center for Structural Genomics, which includes so far about 20 prophage proteins encoded in bacterial genomes.

Distributed structure determination at the JCSG.

The Joint Center for Structural Genomics (JCSG), one of four large-scale structure-determination centers funded by the US Protein Structure Initiative (PSI) through the National Institute for General Medical Sciences, has been operating an automated distributed structure-solution pipeline, Xsolve, for well over half a decade. During PSI-2, Xsolve solved, traced and partially refined 90% of the JCSG's nearly 770 MAD/SAD structures at an average resolution of about 2 Å without human intervention. Xsolve executes many well established publicly available crystallography software programs in parallel on a commodity Linux cluster, resulting in multiple traces for any given target. Additional software programs have been developed and integrated into Xsolve to further minimize human effort in structure refinement. Consensus-Modeler exploits complementarities in traces from Xsolve to compute a single optimal model for manual refinement. Xpleo is a powerful robotics-inspired algorithm to build missing fragments and qFit automatically identifies and fits alternate conformations.

The Joint Center for Structural Genomics: exploration of the human gut microbiome

For more than a decade, the Joint Center for Structural Genomics (JCSG) [1] has been at the forefront of developing tools and methodologies that allow the application of high-throughput structural biology to a broad range of biological and biomedical investigations. In the previous phases of the National Institutes of Health’s Protein Structure Initiative (PSI; 2000 to 2010) [2], we explored structural coverage of uncharted regions of the protein universe [3], as well as a single organism, allowing complete structural reconstruction of the metabolic network of Thermotoga maritima[4]. In the current phase (PSI: Biology; 2010 to 2015), the JCSG is leveraging its high-throughput platform to explore the structural basis for host-microbe interactions in the human microbiome. The emerging field of metagenomics has been particularly enlightening: the human gut microbiome sequencing projects have already uncovered fascinating new families and expansions of known families for adaptation to this environment. The gut microbiota is dominated by poorly characterized bacterial phyla, which contain an unusually high number of uncharacterized proteins that are largely unstudied. Their influence upon human development, physiology, immunity and nutrition is only starting to surface and is thus an exciting new frontier for structural genomics, where we can structurally investigate the contributions of these microorganisms to human health and disease. The JCSG is located at The Scripps Research Institute, the Genomics Institute of the Novartis Research Foundation, University of California at San Diego, the Sanford-Burnham Medical Research Institute and SSRL/Stanford University.

The Joint Center for Structural Genomics: exploration of the human gut microbiome

The Joint Center for Structural Genomics: exploration of the human gut microbiome

Publishing structural genomics results: the JCSG Special Issue

In the last decade we have witnessed the advent of structural genomics initiatives around the world. While traditional hypothesis-driven research continues to produce a wealth of new macromolecular structures, the contribution of structural genomics structures to the Protein Data Bank (PDB) has become ever more important. For example, the USA-based Protein Structure Initiative (PSI) alone has deposited more than 5000 structures with the PDB during phases I and II of the project. While the various structural genomics initiatives have been immensely successful in producing structures, they have found publishing their results to be yet another bottleneck to overcome. Presently, many thousands of structures in the PDB have no publication associated with them at all. When Acta Crystallographica Section F was founded, one of its mandates was to provide an efficient and effective medium for presenting structural genomics results. We have sought new ways to ease the publication bottleneck and, with the help and advice of structural genomics leaders, now offer a platform to showcase these structures, and the experiments upon which they are based, to the wider structural and scientific communities. The key feature is the collection of the results produced by an individual structural genomics effort into a special section, allowing the structural genomics group to organize and annotate their structures and put groups of structures into context. The first example was published in December 2009 and contained eight papers from the RIKEN–UK structural genomics consortium in a special section. We have now completed the ultimate expression of this strategem, our first structural genomics special issue of Acta Crystallographica Section F, published online and entirely open access at http://journals.iucr.org/f/issues/2010/10/00/issconts.html. The Joint Center for Structural Genomics (JCSG), headed by Ian Wilson at the Scripps Institute (La Jolla, USA), contributed 35 fine papers to this issue and the center deserves our thanks for this concerted effort and perseverance. The October 2010 publication of the JCSG Special Issue defines the successful conclusion of an evolving process that started about two years ago during the 2008 IUCr Congress in Osaka, and it marks an important milestone for our journal – the first issue to focus exclusively on structures from an individual structural genomics group We have so far received a lot of positive feedback about this special issue from readers and from the structural genomics community as a whole. However, it is no more than a start, albeit clearly a successful one. We anticipate more special issues and special sections in the future and we already have new requests and proposals under consideration. Finally, we would like to thank our reviewers for their extraordinary generosity of time and energy in refereeing the papers in this special issue.

TOPSAN: use of a collaborative environment for annotating, analyzing and disseminating data on JCSG and PSI structures.

The NIH Protein Structure Initiative centers, such as the Joint Center for Structural Genomics (JCSG), have developed highly efficient technological platforms that are capable of experimentally determining the three-dimensional structures of hundreds of proteins per year. However, the overwhelming majority of the almost 5000 protein structures determined by these centers have yet to be described in the peer-reviewed literature. In a high-throughput structural genomics environment, the process of structure determination occurs independently of any associated experimental characterization of function, which creates a challenge for the annotation and analysis of structures and the publication of these results. This challenge has been addressed by developing TOPSAN (`The Open Protein Structure Annotation Network'), which enables the generation of knowledge via collaborations among globally distributed contributors supported by automated amalgamation of available information. TOPSAN currently provides annotations for all protein structures determined by the JCSG in addition to preliminary annotations on a large number of structures from the other PSI production centers. TOPSAN-enabled collaborations have resulted in insightful structure-function analysis for many proteins and have led to numerous peer-reviewed publications, as exemplified by the articles included in this issue of Acta Crystallographica Section F.

NMR in a crystallography-based high-throughput protein structure-determination environment

An introduction is provided to three papers which compare corresponding protein crystal and NMR solution structures determined by the Joint Center for Structural Genomics (JCSG). Special mention is made of the JCSG strategy for combined use of the two techniques, and of potential applications of the concept of `reference crystal structures', which is introduced in the following three papers.

Structural classification of proteins and structural genomics: new insights into protein folding and evolution.

During the past decade, the Protein Structure Initiative (PSI) centres have become major contributors of new families, superfamilies and folds to the Structural Classification of Proteins (SCOP) database. The PSI results have increased the diversity of protein structural space and accelerated our understanding of it. This review article surveys a selection of protein structures determined by the Joint Center for Structural Genomics (JCSG). It presents previously undescribed β-sheet architectures such as the double barrel and spiral β-roll and discusses new examples of unusual topologies and peculiar structural features observed in proteins characterized by the JCSG and other Structural Genomics centres.