journal menu
abstracts
The Center for Structural Genomics for Infectious Diseases (CSGID) applies structural genomics approaches to biomedically relevant proteins from human pathogens and provides the infectious disease community with a high throughput pipeline for structure determination. Target proteins include drug targets, essential enzymes, virulence factors and vaccine candidates. Bacterial species generally have many acetyl-coenzyme A dependent GCN5-like Acetyl Transferases (GNATs), however, the substrates of most of them are unknown. Proteomic analysis has also revealed extensive post-translational modification of bacterial proteins, especially acetylation of lysine Nε. These observations led the CSGID to develop a high throughput substrate screen and initiate characterization of bacterial GNATs. One of the bacterial GNATs that acetylates lysine residues, is the Pseudomonas aeruginosa protein PA4794, that acetylates both peptides having a C-terminal lysine and the drug, chloramphenicol. Surprisingly, the acetylation of these two substrates by PA4794 is catalyzed by the enzyme using different active site residues and different kinetic mechanisms. Although it was expected that the GNATs would play a major role in protein acetylation, much of the lysine acetylation observed in bacteria is actually due to the metabolite acetylphosphate (1,2). Crystal structures and proteomics experiments revealed what makes some lysine residues particularly sensitive to acetylphosphate dependent lysine acetylation and what is required for subsequent enzymatic deacetylation. CSGID is funded with Federal funds from the National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services, under Contracts No. HHSN272200700058C and HHSN272201200026C and Midwest Center for Structural Genomics by grant GM094585
abstracts
The LabDB laboratory information management system (LIMS) tracks, organizes and analyzes data from chemical and solution management, protein production, crystallization, diffraction, structure solution, and in vitro biochemical and biophysical experiments. The system is comprised of multiple modules specialized for different tasks, such as the Xtaldb system for crystallization or the hkldb module of the HKL-3000 suite for diffraction data collection and structure solution. The biochemical/biophysical experiments tracked by LabDB include spectrophotometric binding and kinetics, thermal shift binding, isothermal titration calorimetry (ITC) and protein quantitation. These tools associate functional and structural experiments, for example, for selecting likely substrates for co-crystallization and soaking experiments. Whenever possible, the system harvests data with no or minimal user intervention from laboratory hardware. Devices that may connect to or import data into LabDB include crystal observation (Rigaku Minstrel HT and Formulatrix Rock Imager), liquid handling (Formulatrix Rock Maker and Emerald Opti-Matrix Maker), chromatography (GE Healthcare AKTA), quantitation (Caliper LabChip GX II and Bio-Rad Gel Doc EZ), RT-PCR (Applied Biosystems 7900HT and Bio-Rad C1000/CFX96) and ITC (MicroCal iTC-200) systems. LabDB is used by two high-throughput PSI:Biology centers (MCSG and NYSGRC) as well as other major NIH consortia (the Center for Structural Genomics of Infectious Diseases and the Enzyme Function Initiative), and track millions of experiments on tens of thousands of targets.[1] The system also provides extensive data mining and analysis tools for translating raw experimental data into information and knowledge. We present examples of analyses generated by the system useful in designing new experiments.
abstracts
"The Structural Biology Knowledgebase (SBKB, http://sbkb.org) was established as a data aggregator to facilitate research design and analysis for a wide variety of biological systems. It serves as a single resource that integrates structure, sequence, and functional annotations plus technical information regarding protein production and structure determination. Researchers can search the SBKB by sequence, PDB ID or UniProt accession code, and receive an up-to-the-minute list of matching 3D experimental structures from the Protein Data Bank, pre-built theoretical models from the Protein Model Portal, annotations from 100+ open biological resources, structural genomics target histories and protocols from TargetTrack, and ready-to-use DNA clones from DNASU. It also possible to find structures according to functional relevance (KB-Rank tool), or find related technologies and publications from the PSI Technology and Publications Portals, respectively. Interactive tools such as real-time theoretical modeling and biophysical parameter prediction also enhance understanding of proteins that are not yet well characterized. Experimentally-focused ""hubs"" collect links to helpful tools and resources for the areas of Structural Targets; Structure, Sequence and Function; Homology Models, Methods and Technologies, and Membrane Proteins. In partnership with the Nature Publishing Group, latest research highlights and articles on specific biological systems are written monthly to share the impact of structural biology. This presentation will demonstrate how the SBKB turns data into knowledge and enables further research. SBKB is funded by a grant from the National Institute of General Medical Sciences of the National Institutes of Health (U01 GM093324)."
abstracts
Metals play vital roles in both the mechanism and architecture of biological macromolecules, and are the most frequently encountered ligands (i.e. non-solvent heterogeneous chemical atoms) in the determination of macromolecular crystal structures. However, metal coordinating environments in protein structures are not always easy to check in routine validation procedures, resulting in an abundance of misidentified and/or suboptimally modeled metal ions in the Protein Data Bank (PDB). We present a solution to identify these problems in three distinct yet related aspects: (1) coordination chemistry; (2) agreement of experimental B-factors and occupancy; and (3) the composition and motif of the metal binding environment. Due to additional strain introduced by macromolecular backbones, the patterns of coordination of metal binding sites in metal-containing macromolecules are more complex and diverse than those found in inorganic or organometallic chemistry. These complications make a comprehensive library of "permitted" coordination chemistry in protein structures less feasible, and the usage of global parameters such as the bond valence method more practical, in the determination and validation of metal binding environments. Although they are relatively infrequent, there are also cases where the experimental B-factor or occupancy of a metal ion suggests careful examination. We have developed a web-based tool called CheckMyMetal [1](http://csgid.org/csgid/metal_sites/) for the quick validation of metal binding sites. Moreover, the acquired knowledge of the composition and spatial arrangement (motif) of the coordinating atoms around the metal ion may also help in the modeling of metal binding sites in macromolecular structures. All of the studies described herein were performed using the NEIGHBORHOOD SQL database [2], which connects information about all modeled non-solvent heterogeneous chemical motifs in PDB structure by vectors describing all contacts to neighboring residues and atoms. NEIGHBORHOOD has broad applications for the validation and data mining of ligand binding environments in the PDB.
