PrePS - Prenylation Prediction
Suite
Protein CaaX Farnesylation, CaaX Geranylgeranylation and Rab Geranylgeranylation
PRENbase
Database of Prenylated Proteins
Prenylation refers to the posttranslational modification of proteins with isoprenyl anchors. These lipid moieties are typically involved in mediating protein-membrane but also protein-protein interactions of prominent cellular proteins. 3 eukaryotic enzymes are known to catalyze the lipid transfer. The first two, farnesyltransferase (FT) and geranylgeranyltransferase 1 (GGT1), recognize the so-called CaaX-box in the C-termini of substrate proteins and attach farnesyl (15 carbon polyisopren) or geranylgeranyl (20 carbon polyisopren), respectively, to a required and spatially fixed cysteine in that motif. The third enzyme, geranylgeranyltransferase 2 (GGT2 or RabGGT) recognizes the complex of Rab GTPase substrate proteins with a specific Rab escort protein (REP) to attach one or two geranylgeranyl anchors to cysteines in a more flexible but also C-terminal motif.
Literature reviews:
Casey PJ, Seabra MC.
Protein prenyltransferases.
J Biol Chem. 1996 Mar 8;271(10):5289-92.
Sinensky M.
Recent advances in the study of prenylated proteins.
Biochim Biophys Acta. 2000 Apr 12;1484(2-3):93-106.
Roskoski R Jr.
Protein prenylation: a pivotal posttranslational process.
Biochem Biophys Res Commun. 2003 Mar 28;303(1):1-7.
Maurer-Stroh S, Washietl S, Eisenhaber F.
Protein prenyltransferases.
Genome Biol. 2003;4(4):212. Epub 2003 Apr 01.
PrePS stands for Prenylation Prediction Suite and combines three predictors for protein CaaX farnesylation, CaaX geranylgeranylation and Rab geranylgeranylation in one webinterface. The predictors aim to model the substrate-enzyme interactions based on refinement of the recognition motifs for each of the prenyltransferases. Motif information has been extracted from sets of known substrates (learning sets) and specific scoring functions have been created utilizing both sequence as well as physical property profiles including interpositional correlations to account for partially overlapping substrate specificities. The PrePS selectively assigns the modifying enzyme to predicted substrate proteins and sensitively filters out false positive predictions based on the methodology that has already been applied successfully for the prediction of GPI-anchors, myristoylation and PTS1 peroxisomal targeting.
Simply copy/paste your sequence in the input form in single letter amino acid code or FASTA format.
What are the provided options?
You can select the prenylation of which of the three enzymes you want to be predicted.
The option for evOluation over NCBI's non-redundant database (NR) allows for
evaluating the evolutionary motif conservation (evOluation). When selected, this option initiates a BLAST search for
homologues in NR which takes, depending on our server load, between 0.5 and 2 minutes. Then, the BLAST hits are
automatically submitted to PrePS and the prenylation prediction results annotated in the C-terminal alignment of the
homologues.
The option for evOluation over PRENbase will initiate a BLAST against PRENbase.
The BLAST hits of your sequence in PRENbase are arranged into their respective clusters and ranked according to the
best E-value with any cluster member. The output format of the clusters is very similar to the standard PRENbase (see
here for explanation of the format). The "Explore" view leads to the individual hitting
sequences with the E-value added to the default motif and prediction information.
How do I interpret PrePS results?
The two CaaX prenylation predictors (FT and GGT1) provide a score and estimation of the probability of false positive prediction, while the GGT2 predictor gives scores and E-values of the HMMer searches. Details of the profile and physical property terms of the scoring function or the HMMer alignments, respectively, are provided (follow link). Penalties on specific positions or regions can also be used to rationalize whether and why certain query sequences or artificial constructs (e.g. intended for membrane targeting) might be less suitable prenylation targets.
Please keep in mind that there are overlapping substrate specificities between the three prenyltransferases. As a simple rule, if a protein is a predicted GGT2 substrate then it often is geranylgeranylated by GGT2 in vivo also when it could be alternatively modified by FT or GGT1 in vitro (as would be indicated by a prediction as FT or GGT1 substrate). In the case of predicted substrates for both FT and GGT1, our method was found to correlate best with experimentally verified enzyme affinities when comparing the relative scores of the predictors with the GGT1 score divided by 3 (because GGT1 seems to have a 3-fold lower activity compared to FT). Proteins ending with a -CxxL motif were previously thought to be classical GGT1 substrates but can also be alternatively modified to a certain extent by FT in vitro due to its broad substrate specificity. In vivo, however, these proteins ending with a leucine are likely to be geranylgeranylated by GGT1.
PrePS gives additional quality assignments for better comparison of the individual predictions. These are, with decreasing likelihood of being a prenylation target: +++, ++, +, -, -- and ---. The different PLUS attributes describe the quality of proteins as PREDICTED prenylation substrates. A MINUS attribute means that the query sequence is NOT PREDICTED to be prenylated with distinction on how far the query has been from the prediction limit.
In case the evOluation option was used, the evolutionary motif conservation can be evaluated from the alignment of the C-termini of homologous sequences in combination with the respective PrePS predictions. For example, a prenylation motif that is conserved in a series of homologues in different organisms might be biologically more important than an isolated prediction.
IMPORTANT: Please note that PrePS only can tell you whether a protein would be processed by the prenylating enzymes when provided as substrate. This prediction does not necessarily imply a cellular context for the query protein that would allow in vivo access to the respective enzymes.
The manuscript describing PrePS is available from the Genome
Biology website (http://genomebiology.com/2005/6/6/R55) and can
be cited as:
Maurer-Stroh S, Eisenhaber F
Refinement and prediction of protein prenylation motifs
Genome Biology 2005, 6:R55 doi:10.1186/gb-2005-6-6-r55
PRENbase is an annotated database of known and predicted prenylated
proteins (predicted using the PrePS - Prenylation Prediction Suite). Homologous proteins
are merged into clusters. A search interface allows sophisticated queries for the experimental status of the
modification (known/predicted...), exclusive or shared types of modifying enzymes (FT, GGT1, GGT2) as well as for
evolutionary conservation by constraining the taxonomic distribution within these clusters or for single
sequences.
HumanPRENbase is a derivate of PRENbase with focus on human prenylated
proteins. In PRENbase, paralogous proteins are clustered together in their respective larger family when they are
highly similar to each other. This happens, for example, with the different Ras proteins H-Ras, K-Ras, N-ras, etc.
Paralogues can nevertheless have different functions or often similar but more specialized roles. Especially in the
light of the importance of prenylation of several members of the Ras, Rab and Rho GTPase families, we sought a
solution to investigate the prenylation status of individual human proteins rather than larger families they are part
of. Therefore, we employed a scheme of reciprocal BLASTs in order to identify the true orthologues of a set of
prenylated human proteins (see accompanying publication for details). 238 individual human proteins and their
orthologues form the clusters in HumanPRENbase.
Do I have to understand the search interface to use PRENbase?
No. The default settings will give access to the collection of both known and predicted eukaryotic and viral prenylated proteins, which can then be browsed. Below the interface we have listed a series of standard queries that you might find useful or are particularly biologically important.
Tutorial for customized searches.
The search interface is designed to allow sophisticated queries for the
experimental status of the modification (known/predicted...), exclusive or shared types of modifying enzymes (FT,
GGT1, GGT2) as well as for evolutionary conservation by constraining the taxonomic distribution within these clusters
or for single sequences.
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