Georg Schneider

SIFT web server: predicting effects of amino acid substitutions on proteins

The Sorting Intolerant from Tolerant (SIFT) algorithm predicts the effect of coding variants on protein function. It was first introduced in 2001, with a corresponding website that provides users with predictions on their variants. Since its release, SIFT has become one of the standard tools for characterizing missense variation. We have updated SIFT’s genome-wide prediction tool since our last publication in 2009, and added new features to the insertion/deletion (indel) tool. We also show accuracy metrics on independent data sets. The original developers have hosted the SIFT web server at FHCRC, JCVI and the web server is currently located at BII. The URL is http://sift-dna.org (24 May 2012, date last accessed).

Towards complete sets of farnesylated and geranylgeranylated proteins

Three different prenyltransferases attach isoprenyl anchors to C-terminal motifs in substrate proteins. These lipid anchors serve for membrane attachment or protein–protein interactions in many pathways. Although well-tolerated selective prenyltransferase inhibitors are clinically available, their mode of action remains unclear since the known substrate sets of the various prenyltransferases are incomplete. The Prenylation Prediction Suite (PrePS) has been applied for large-scale predictions of prenylated proteins. To prioritize targets for experimental verification, we rank the predictions by their functional importance estimated by evolutionary conservation of the prenylation motifs within protein families. The ranked lists of predictions are accessible as PRENbase (http://mendel.imp.univie.ac.at/sat/PrePS/PRENbase) and can be queried for verification status, type of modifying enzymes (anchor type), and taxonomic distribution. Our results highlight a large group of plant metal-binding chaperones as well as several newly predicted proteins involved in ubiquitin-mediated protein degradation, enriching the known functional repertoire of prenylated proteins. Furthermore, we identify two possibly prenylated proteins in Mimivirus. The section HumanPRENbase provides complete lists of predicted prenylated human proteins—for example, the list of farnesyltransferase targets that cannot become substrates of geranylgeranyltransferase 1 and, therefore, are especially affected by farnesyltransferase inhibitors (FTIs) used in cancer and anti-parasite therapy. We report direct experimental evidence verifying the prediction of the human proteins Prickle1, Prickle2, the BRO1 domain–containing FLJ32421 (termed BROFTI), and Rab28 (short isoform) as exclusive farnesyltransferase targets. We introduce PRENbase, a database of large-scale predictions of protein prenylation substrates ranked by evolutionary conservation of the motif. Experimental evidence is presented for the selective farnesylation of targets with an evolutionary conserved modification site.

MYRbase: analysis of genome-wide glycine myristoylation enlarges the functional spectrum of eukaryotic myristoylated proteins

We evaluated the evolutionary conservation of glycine myristoylation within eukaryotic sequences. Our large-scale cross-genome analyses, available as MYRbase, show that the functional spectrum of myristoylated proteins is currently largely underestimated. We give experimental evidence for in vitro myristoylation of selected predictions. Furthermore, we classify five membrane-attachment factors that occur most frequently in combination with, or even replacing, myristoyl anchors, as some protein family examples show.

ANNIE: integrated de novo protein sequence annotation

Function prediction of proteins with computational sequence analysis requires the use of dozens of prediction tools with a bewildering range of input and output formats. Each of these tools focuses on a narrow aspect and researchers are having difficulty obtaining an integrated picture. ANNIE is the result of years of close interaction between computational biologists and computer scientists and automates an essential part of this sequence analytic process. It brings together over 20 function prediction algorithms that have proven sufficiently reliable and indispensable in daily sequence analytic work and are meant to give scientists a quick overview of possible functional assignments of sequence segments in the query proteins. The results are displayed in an integrated manner using an innovative AJAX-based sequence viewer. ANNIE is available online at: http://annie.bii.a-star.edu.sg. This website is free and open to all users and there is no login requirement.

Application of a sensitive collection heuristic for very large protein families: Evolutionary relationship between adipose triglyceride lipase (ATGL) and classic mammalian lipases

BackgroundManually finding subtle yet statistically significant links to distantly related homologues becomes practically impossible for very populated protein families due to the sheer number of similarity searches to be invoked and analyzed. The unclear evolutionary relationship between classical mammalian lipases and the recently discovered human adipose triglyceride lipase (ATGL; a patatin family member) is an exemplary case for such a problem.ResultsWe describe an unsupervised, sensitive sequence segment collection heuristic suitable for assembling very large protein families. It is based on fan-like expanding, iterative database searches. To prevent inclusion of unrelated hits, additional criteria are introduced: minimal alignment length and overlap with starting sequence segments, finding starting sequences in reciprocal searches, automated filtering for compositional bias and repetitive patterns. This heuristic was implemented as FAMILYSEARCHER in the ANNIE sequence analysis environment and applied to search for protein links between the classical lipase family and the patatin-like group.ConclusionThe FAMILYSEARCHER is an efficient tool for tracing distant evolutionary relationships involving large protein families. Although classical lipases and ATGL have no obvious sequence similarity and differ with regard to fold and catalytic mechanism, homology links detected with FAMILYSEARCHER show that they are evolutionarily related. The conserved sequence parts can be narrowed down to an ancestral core module consisting of three β-strands, one α-helix and a turn containing the typical nucleophilic serine. Moreover, this ancestral module also appears in numerous enzymes with various substrate specificities, but that critically rely on nucleophilic attack mechanisms.

Transmembrane helix: simple or complex

Transmembrane helical segments (TMs) can be classified into two groups of so-called ‘simple’ and ‘complex’ TMs. Whereas the first group represents mere hydrophobic anchors with an overrepresentation of aliphatic hydrophobic residues that are likely attributed to convergent evolution in many cases, the complex ones embody ancestral information and tend to have structural and functional roles beyond just membrane immersion. Hence, the sequence homology concept is not applicable on simple TMs. In practice, these simple TMs can attract statistically significant but evolutionarily unrelated hits during similarity searches (whether through BLAST- or HMM-based approaches). This is especially problematic for membrane proteins that contain both globular segments and TMs. As such, we have developed the transmembrane helix: simple or complex (TMSOC) webserver for the identification of simple and complex TMs. By masking simple TM segments in seed sequences prior to sequence similarity searches, the false-discovery rate decreases without sacrificing sensitivity. Therefore, TMSOC is a novel and necessary sequence analytic tool for both the experimentalists and the computational biology community working on membrane proteins. It is freely accessible at http://tmsoc.bii.a-star.edu.sg or available for download.

Induction of Short Interspersed Nuclear Repeat- containing Transcripts in Epithelial Cells upon Infection with a Chicken Adenovirus

Chicken embryo lethal orphan adenovirus (CELO) is used as a vector for expression of exogenous genes in mammalian cells. Here, we analyzed transcriptional alterations in mouse epithelial host cells following infection with CELO using cDNA microarray analysis. Sequence data characterization revealed that a major portion of CELO-induced genes contained short interspersed nuclear elements of the B2 subclass (B2 SINEs). In fact, we could identify SINEs and other repetitive sequences as contributing significantly to the cDNAs used for microarray construction. Moreover, we found that the CELO protein Gam1 was able to mediate transcriptional activation of these B2 SINE-containing RNAs. We hypothesize that upregulation of B2-SINE-containing RNAs could be a novel contribution of Gam1 to CELO host cell infection.

DOUTfinder—identification of distant domain outliers using subsignificant sequence similarity

DOUTfinder is a web-based tool facilitating protein domain detection among related protein sequences in the twilight zone of sequence similarity. The sequence set required for this analysis can be provided by the user or will be collected using PSI-BLAST if a single sequence is given as an input. The obtained sequence family is analyzed for known Pfam and SMART domains, and the thereby identified subsignificant domain similarities are evaluated further. Domains with several subthreshold hits in the query set are ranked based on a sum-score function and likely homologous domains are suggested according to established cut-offs. By providing a post-filtering procedure for subsignificant domain hits DOUTfinder allows the detection of non-trivial domain relationships and can thereby lead to new insights into the function and evolution of distantly related sequence families. DOUTfinder is available at .

Protein Sequence–Structure–Function–Network Links Discovered with the ANNOTATOR Software Suite: Application to ELYS/Mel-28

While very little genomic sequence is interpretable in terms of biological mechanism directly, the chances are much better for protein-coding genes that can be translated into protein sequences. This review considers the different concepts applicable to sequence analysis and function prediction of globular and non-globular protein segments. The publicly accessible ANNOTATOR software environment integrates most of the reliable protein sequence-based function prediction methods, protein domain databases and pathway, and protein–protein interaction collections developed in academia. As application example, the structural and functional domains of mel-28/ELYS, an important nuclear protein, are delineated and are proposed for experimental follow-up in structural biology and functional studies.