Workshop on Molecular Animation Bromberg, Sarina; Chiu, Wah; Ferrin, Thomas E.
Structure (London),
10/2010, Volume:
18, Issue:
10
Journal Article, Conference Proceeding
Peer reviewed
Open access
From February 25 to 26, 2010, in San Francisco, the Resource for Biocomputing, Visualization, and Informatics (RBVI) and the National Center for Macromolecular Imaging (NCMI) hosted a molecular ...animation workshop for 21 structural biologists, molecular animators, and creators of molecular visualization software. Molecular animation aims to visualize scientific understanding of biomolecular processes and structures. The primary goal of the workshop was to identify the necessary tools for producing high-quality molecular animations, understanding complex molecular and cellular structures, creating publication supplementary materials and conference presentations, and teaching science to students and the public. Another use of molecular animation emerged in the workshop: helping to focus scientific inquiry about the motions of molecules and enhancing informal communication within and between laboratories.
MODBASE (http://salilab.org/modbase) is a relational database of annotated comparative protein structure models for all available protein sequences matched to at least one known protein structure. ...The models are calculated by MODPIPE, an automated modeling pipeline that relies on the MODELLER package for fold assignment, sequence–structure alignment, model building and model assessment (http:/salilab.org/modeller). MODBASE uses the MySQL relational database management system for flexible querying and CHIMERA for viewing the sequences and structures (http://www.cgl.ucsf.edu/chimera/). MODBASE is updated regularly to reflect the growth in protein sequence and structure databases, as well as improvements in the software for calculating the models. For ease of access, MODBASE is organized into different data sets. The largest data set contains 1 262 629 models for domains in 659 495 out of 1 182 126 unique protein sequences in the complete Swiss‐Prot/TrEMBL database (August 25, 2003); only models based on alignments with significant similarity scores and models assessed to have the correct fold despite insignificant alignments are included. Another model data set supports target selection and structure‐based annotation by the New York Structural Genomics Research Consortium; e.g. the 53 new structures produced by the consortium allowed us to characterize structurally 24 113 sequences. MODBASE also contains binding site predictions for small ligands and a set of predicted interactions between pairs of modeled sequences from the same genome. Our other resources associated with MODBASE include a comprehensive database of multiple protein structure alignments (DBALI, http://salilab.org/dbali) as well as web servers for automated comparative modeling with MODPIPE (MODWEB, http://salilab. org/modweb), modeling of loops in protein structures (MODLOOP, http://salilab.org/modloop) and predicting functional consequences of single nucleotide polymorphisms (SNPWEB, http://salilab. org/snpweb).
structureViz is a Cytoscape plug-in that links the visualization of biological networks provided by Cytoscape with the visualization and analysis of macromolecular structures and sequences provided ...by UCSF Chimera. When combined with Cytoscape and Chimera, structureViz provides the first tool that links these two critical aspects of computational analysis in a straightforward manner. structureViz includes commands to open structures in Chimera and align them using Chimera's sequence-structure analysis tools. When a structure is opened, structureViz provides an alternative interface to Chimera: the Cytoscape Molecular Structure Navigator. This interface uses a tree-based paradigm to allow users to select and affect the display of models, chains and residues, mostly through the use of context menus. Contact: scooter@cgl.ucsf.edu
Systemic carnitine deficiency (SCD) is a rare autosomal recessive disease resulting from defects in the OCTN2 ( SLC22A5 ) gene, which encodes the high-affinity plasma membrane carnitine transporter. ...Although OCTN2 is fairly well studied in its
relationship with SCD, little is known about the carrier frequency of disease-causing alleles of OCTN2, or of more common
functional polymorphisms in this gene. To address these issues, we screened for genetic variants in the OCTN2 coding region
by direct sequencing of the exons and flanking intronic region of OCTN2 in a large sample ( n = 276) of ethnically diverse subjects. In addition, we established lymphoblastoid cell lines from subjects homozygous for
either allele of the previously identified promoter region variant, -207G>C. We found eight amino acid sequence variants of
OCTN2, of which three (Phe17Leu, Leu144Phe, and Pro549Ser) were polymorphic in at least one ethnic group. When assayed for
functional activity by expression in human embryonic kidney 293 cells, using as probes both the endogenous substrate ( l -carnitine) and the organic cation tetraethylammonium, three variants showed functional differences from the reference OCTN2
(Phe17Leu, Tyr449Asp, Val481Phe; p < 0.05). Further studies of the Phe17Leu polymorphism showed a reduced V max for l -carnitine transport to approximately 50% of the reference OCTN2. Confocal microscopy studies using an OCTN2-GFP fusion protein
showed that Phe17Leu had distinct subcellular localization from the reference OCTN2, with diffuse cytoplasmic retention of
Phe17Leu, in contrast to reference OCTN2, which localized specifically to the plasma membrane. Lymphoblasts from subjects
homozygous for the -207G allele showed increased l -carnitine transport compared with the -207C/C homozygotes ( p < 0.05). This study suggests that although loss-of-function mutations in OCTN2 are likely to be rare, common variants of
OCTN2 found in healthy populations may contribute to variation in the disposition of carnitine and some clinically used drugs.
Apical reabsorption from the urine has been shown to be important for such processes as the maintenance of critical metabolites in the blood and the excretion of nephrotoxic compounds. The solute ...carrier (SLC) transporter OAT4 (SLC22A11) is expressed on the apical membrane of renal proximal tubule cells and is known to mediate the transport of a variety of xenobiotic and endogenous organic anions. Functional characterization of genetic variants of apical transporters thought to mediate reabsorption, such as OAT4, may provide insight into the genetic factors influencing the complex pathways involved in drug elimination and metabolite reclamation occurring in the kidney. Naturally occurring genetic variants of OAT4 were identified in public databases and by resequencing DNA samples from 272 individuals comprising 4 distinct ethnic groups. Nine total nonsynonymous variants were identified and functionally assessed using uptake of three radiolabeled substrates. A nonsense variant, R48Stop, and three other variants (R121C, V155G, and V155M) were found at frequencies of at least 2% in an ethnic group specific fashion. The L29P, R48Stop, and H469R variants displayed a complete loss of function, and kinetic analysis identified a reduced V(max) in the common nonsynonymous variants. Plasma membrane levels of OAT4 protein were absent or reduced in the nonfunctional variants, providing a mechanistic reason for the observed loss of function. Characterization of the genetic variants of reabsorptive transporters such as OAT4 is an important step in understanding variability in tubular reabsorption with important implications in innate homeostatic processes and drug disposition.
With the increase in the number of large, 3D, high-resolution nucleic acid structures, particularly of the 30S and 50S ribosomal subunits and the intact bacterial ribosome, advancements in the ...visualization of nucleic acid structural features are essential. Large molecular structures are complicated and detailed, and one goal of visualization software is to allow the user to simplify the display of some features and accent others. We describe an extension to the UCSF Chimera molecular visualization system for the purpose of displaying and highlighting nucleic acid characteristics, including a new representation of sugar pucker, several options for abstraction of base geometries that emphasize stacking and base pairing, and an adaptation of the ribbon backbone to accommodate the nucleic acid backbone. Molecules are displayed and manipulated interactively, allowing the user to change the representations as desired for small molecules, proteins and nucleic acids. This software is available as part of the UCSF Chimera molecular visualization system and thus is integrated with a suite of existing tools for molecular graphics.
BACKGROUNDOCTN1 is a multispecific transporter of organic cations and zwitterions, including several clinically important drugs as well as the antioxidant ergothioneine. OCTN1 is highly expressed in ...the kidney, where it is thought to aid in active secretion of organic cations, and may facilitate the active reabsorption of ergothioneine. Genetic variation in OCTN1 may help to explain interindividual variability in the pharmacokinetics of many cationic or zwitterionic drugs.
METHODSWe screened for human genetic variants in the OCTN1 coding region by direct sequencing in a large sample (n=270) of ethnically diverse healthy volunteers.
RESULTSSix protein sequence-altering variants were identified, including five-amino-acid substitutions and one nonsense mutation. Two of the variants, T306I and L503F, were polymorphic, occurring at frequencies of 37 and 19%, respectively, in the total sample. Allele frequencies are varied by ethnicity. In biochemical assays, two of the variants (D165G and R282X) resulted in complete loss of transport function, and one variant (M205I) caused a reduction in activity to approximately 50% of the reference sequence protein. One variant, L503F, showed altered substrate specificity; this variant occurred at particularly high allele frequency (42%) in the European-American participants in our sample. Subcellular localization and ergothioneine inhibition kinetics were similar among the common amino-acid sequence variants of OCTN1.
CONCLUSIONSThe common OCTN1-L503F variant may explain a significant amount of population variation in the pharmacokinetics of OCTN1 substrate drugs. The rare loss-of-function variants provide a rational tool for studying the importance of ergothioneine in humans in vivo.
The BayGenomics gene-trap resource (http://baygenomics.ucsf.edu) provides researchers with access to thousands of mouse embryonic stem (ES) cell lines harboring characterized insertional mutations in ...both known and novel genes. Each cell line contains an insertional mutation in a specific gene. The identity of the gene that has been interrupted can be determined from a DNA sequence tag. Approximately 75% of our cell lines contain insertional mutations in known mouse genes or genes that share strong sequence similarities with genes that have been identified in other organisms. These cell lines readily transmit the mutation to the germline of mice and many mutant lines of mice have already been generated from this resource. BayGenomics provides facile access to our entire database, including sequence tags for each mutant ES cell line, through the World Wide Web. Investigators can browse our resource, search for specific entries, download any portion of our database and BLAST sequences of interest against our entire set of cell line sequence tags. They can then obtain the mutant ES cell line for the purpose of generating knockout mice.
The concentrative nucleoside transporter, CNT1 (SLC28A1), mediates the cellular uptake of naturally occurring pyrimidine nucleosides
and many structurally diverse anticancer and antiviral nucleoside ...analogs. As a first step toward understanding whether genetic
variation in CNT1 contributes to variation in the uptake and disposition of clinically used nucleoside analogs, we determined
the haplotype structure and functionally analyzed all coding region variants of CNT1 identified in ethnically diverse populations
(100 African Americans, 100 European Americans, 30 Asians, 10 Mexican Americans, and 7 Pacific Islanders) ( Leabman et al., 2003 ). A total of 58 coding region haplotypes were identified using PHASE analysis, 44 of which contained at least one amino acid
variant. More than half of the coding region haplotypes were population-specific. Using site-directed mutagenesis, 15 protein-altering
CNT1 variants, including one amino acid insertion and one base pair (bp) deletion, were constructed and expressed in Xenopus laevis oocytes. All variant transporters took up 3 Hthymidine with the exception of CNT1-Ser546Pro, a rare variant, and CNT1â1153del, a single bp deletion found at a frequency
of 3% in the African American population. The bp deletion results in a frame-shift followed by a stop-codon. The anticancer
nucleoside analog gemcitabine had a reduced affinity for CNT1-Val189Ile (a common CNT1 variant found at a frequency of 26%)
compared with reference CNT1 (IC 50 = 13.8 ± 0.60 μM for CNT1-reference and 23.3 ± 1.5 μM for CNT1-Val189Ile, p < 0.05). These data suggest that common genetic variants of CNT1 may contribute to variation in systemic and intracellular
levels of anti-cancer nucleoside analogs.
CyAnimator (http://apps.cytoscape.org/apps/cyanimator) is a Cytoscape app that provides a tool for simple animations of Cytoscape networks. The tool allows you to take a series of snapshots ...(CyAnimator calls them frames) of Cytoscape networks. For example, the first frame might be of a network shown from a ”zoomed out” viewpoint and the second frame might focus on a specific group of nodes. Once these two frames are captured by the tool, it can animate between them by interpolating the changes in location, zoom, node color, node size, edge thickness, presence or absence of annotations, etc. The animations may be saved as a series of individual frames, animated GIFs, or H.264/MP4 movies.
CyAnimator is available from within the Cytoscape App Manager or from the Cytoscape app store.
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