truncation of protein sequences for fast profile alignment with application to subcellular...
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Truncation of Protein Sequences for Fast Profile Alignment with Application
to Subcellular Localization
Man-Wai MAK and Wei WANG The Hong Kong Polytechnic University
Sun-Yuan KUNGPrinceton University
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Contents
1. Introduction– Cell Organelles and Proteins Subcellular Localization– Signal-Based vs. Homology-Based Methods
2. Speeding Up the Prediction Process– Predicting Cleaving Site Location– Truncating Profiles vs. Truncating Sequences– Perturbational Discriminant Analysis
3. Experiments and Results4. Conclusions
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Organelles• Cells have a set of organelles that are specialized for carrying out
one or more vital functions.• Proteins must be transported to the correct organelles of a cell to
properly perform their functions. • Therefore, knowing the subcellular localization is one step towards
understanding the functions of proteins.
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Proteins and Their Subcellular Location
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Subcellular Localization Prediction
Two key methods:1. Signal-based2. Homology-based
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Signal-Based Method
Source: S. R. Goodman, Medical Cell Biology, Elsevier, 2008.
• The amino acid sequence of a protein contains information about its organelle destination.
• Typically, the information can be found within a short segment of 20 to 100 amino acids preceding the cleavage site.
• Signal-based methods (e.g. TargetP) can determine the cleavage site location
Cleavage site
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Full-length Query Sequence
S(1)=KNKA···S(2)=KAKN···
··
S(N)=KGLL···
Full-length Training sequences
Align with each of the training
sequences ...
SVM classifier
N-dim alignment vector
Subcellular Location
1
N
Advantage:• Can predict sequences that do
not have cleavage sites.Drawback:• Given a query sequence, we
need to align it with every training sequence in the training set, causing long computation time.
Homology-Based Method
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Sequences Length Distribution
• Many sequences are fairly long, thus, aligning the whole sequence will take long computation time.
• cTP, mTP and SP are under 100 AAs only and contain the most relevant segment.
• Computation saving can be achieved by aligning the signal segments only.
Length distribution of Seq.
Sequence Length
SP
820
Ext:
Mit:
Chl:
35
mTP
1050
18
cTP
760
Cleavage Site
Cleavage Site
Cleavage Site
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Proposed Method: Aligning the Segments that Contain the Most Relevant Info.
Signal-based Cleavage Site Predictor(e.g. TargetP)
N
truncate
Homology-based Method
SubcellularLocation
CAmino Acid Sequence…
Truncated sequence
Cleavage Site
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Aligning Profiles Vs. Aligning Sequences
shortprofilesequence
shortCut SVM or
KPDAPairwise
AlignmentPSI-
BLASTSubcellular
Location
ScoreVector
shortprofileprofile
LongPSI-BLAST
SVM or KPDA
PairwiseAlignmentCut
SubcellularLocation
ScoreVector
Scheme I
Scheme II
shortprofilesequence
shortCut SVM or
KPDAPairwise
AlignmentPSI-
BLASTSubcellular
Location
ScoreVector
shortprofileprofile
LongPSI-BLAST
SVM or KPDA
PairwiseAlignmentCut
SubcellularLocation
ScoreVector
Scheme I
Scheme II
QuerySequence
Scheme I : Truncate the profilesScheme II : Truncate the sequences
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Perturbational Discriminant Analysis
Input Space Hilbert Space
Input and Hilbert Spaces:
Empirical Space:
Nxk )(),( 1 xxK
Empirical Space
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Perturbational Discriminant Analysis
• The objective of PDA is to find an optimal discriminant function in the Hilbert space or empirical space:
• The optimal solution (see derivation in paper) in the empirical space is
• ρ represents the noise (uncertainty) level in the measurement. It also ensures numerical stability of the matrix inverse.
• Ρ = 1 in this work.
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Perturbational Discriminant Analysis
3 classes of 2-dim data in the input space
RBF kernal matrix K
Projection onto the 2-dim PDA space
Decision boundaries in the input space
Example on 2-D Data
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Perturbational Discriminant Analysis
Application to Sequence Classification
Training sequences
PSI-BLASTPairwise
AlignmentComputePDA Para
TrainingProfiles K
Test sequence
PSI-BLASTAlign withTraining Profiles
ComputePDA Score
TestProfile
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Perturbational Discriminant AnalysisApplication to Multi-Class Problems
1-vs-Rest PDA Classifier:
)(1 xf )(xfC
MAXNET
)(2 xf
x
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Perturbational Discriminant Analysis
Application to Multi-Class Problems
Cascaded PDA-SVM Classifier:
Test sequence Project onto
(C–1)-dimPDA space
1-vs-restSVM
Classifier
Class label
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Experiments
Materials:
• Eukaryotic sequences extracted from Swiss-Prot 57.5• Ext, Mit, and Chl contain experimentally determined cleavage sites• 25% Sequence identity (based on BLASTclust)
Performance Evaluation:
• 5-Fold cross validation• Prediction accuracy and Matthew’s correlation coefficient (MCC)
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shortprofilesequence
shortCut SVM or
KPDAPairwise
AlignmentPSI-
BLASTSubcellular
Location
ScoreVector
shortprofileprofile
LongPSI-BLAST
SVM or KPDA
PairwiseAlignmentCut
SubcellularLocation
ScoreVector
Scheme I
Scheme II
shortprofilesequence
shortCut SVM or
KPDAPairwise
AlignmentPSI-
BLASTSubcellular
Location
ScoreVector
shortprofileprofile
LongPSI-BLAST
SVM or KPDA
PairwiseAlignmentCut
SubcellularLocation
ScoreVector
Scheme I
Scheme II
QuerySequence
Kernel matrix(Scheme I)
Kernel matrix(Scheme II)
Comparing Kernel Matrices
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Sensitivity Analysis
• The localization performance degrades when the cut-off position drifts away from the ground-truth cleavage site.
• mTP and cTP are more sensitive
to the error of cleavage site prediction than Ext.
19Cut-off Position
p-16 p-8 p-2 p p+2 p+16 p+32 p+64
Ground-truth cleavage site
Cyt/Nuc
Overall
Mit
Chl
Ext
Cut Seq. at p±xp: gournd-truth cleave site
Subcellular localization(PairProSVM)
Subcellular location
Seq
Sub
cellu
lar
Loca
liatio
n A
ccur
acy
(%)
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Performance of Cleavage Site Prediction
• Conditional Random Field (CRF) is better than TargetP(Plant) in terms of predicting the cleavage sites of signal peptide (Ext) but is worse than TargetP(Nonplant).
• CRF is slightly inferior to TargetP in predicting the cleavage sites of mitochondria, but it is significantly better than TargetP in predicting the cleavage site of chloroplasts.
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Targe
tP(P
lant)
Targe
tP(N
onPlan
t)
CRF
Category
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Findings: Profile creation time can be substantially reduced by truncating the protein sequences at the cleavage sites.
Comparing Profile Creation Time
shortprofilesequence
shortCut SVM or
KPDAPairwise
AlignmentPSI-
BLASTSubcellular
Location
ScoreVector
shortprofileprofile
LongPSI-BLAST
SVM or KPDA
PairwiseAlignmentCut
SubcellularLocation
ScoreVector
Scheme I
Scheme II
shortprofilesequence
shortCut SVM or
KPDAPairwise
AlignmentPSI-
BLASTSubcellular
Location
ScoreVector
shortprofileprofile
LongPSI-BLAST
SVM or KPDA
PairwiseAlignmentCut
SubcellularLocation
ScoreVector
Scheme I
Scheme II
QuerySequence
shortprofilesequence
shortCut SVM or
KPDAPairwise
AlignmentPSI-
BLASTSubcellular
Location
ScoreVector
shortprofileprofile
LongPSI-BLAST
SVM or KPDA
PairwiseAlignmentCut
SubcellularLocation
ScoreVector
Scheme I
Scheme II
shortprofilesequence
shortCut SVM or
KPDAPairwise
AlignmentPSI-
BLASTSubcellular
Location
ScoreVector
shortprofileprofile
LongPSI-BLAST
SVM or KPDA
PairwiseAlignmentCut
SubcellularLocation
ScoreVector
Scheme I
Scheme II
QuerySequence
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Findings: The training time of 1-vs-rest PDA and Cascaded PDA-SVM are substantially shorter than that of SVM.
Training and Classification Time
)(1 xf )(xfC
MAXNET
)(2 xf
x
Project onto(C–1)-dim
PDA space
1-vs-restSVM
Classifier
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Findings: In terms of localization accuracy, the proposed “Signal+Homology” method performs slightly better than the signal-based TargetP and is substantially better than the homology-based SubLoc.
Compare with State-of-the-Art Localization Predictors
ConditionalRandomFields
LocalizationAccuracy (%)
Subcellular localization(SubLoc/TargetP)
Cleavage site prediction(TargetP/CRF)
Subcellular localization(PairProSVM)
Subcellularlocation
Queryseq.
Subcellular localization(SubLoc/TargetP)
Cleavage site prediction(TargetP/CRF)
Subcellular localization(PairProSVM)
Subcellularlocation
Queryseq.
MCC
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Conclusion
• Fast subcellular-localization-prediction can be achieved by a cascaded fusion of signal-based and homology-based methods.
• As far as localization accuracy is concerned, it does not matter whether we truncate the sequences or truncate the profiles. However, truncating the sequence can save the profile creation time by 6 folds.
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Compare with State-of-the-Art Localization PredictorsSubcellular localization
(SubLoc/TargetP)
Cleavage site prediction(TargetP/CRF)
Subcellular localization(PairProSVM)
Subcellularlocation
Queryseq.
Subcellular localization(SubLoc/TargetP)
Cleavage site prediction(TargetP/CRF)
Subcellular localization(PairProSVM)
Subcellularlocation
Queryseq.
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Performance of Cascaded Fusion
• The computation time for full-length profile alignment is a striking 116 hours
• Our method not only leads to nearly a 20 folds reduction in computation time but also boosts the prediction performance.
Full-length
Seq.
Seq. with Csite predicted by TargetP(P)
Seq. with Csite predicted by TargetP(N)
Seq. with Csite predicted by CRF
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Time
Subcellular localization accuracy
Acc(%)
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1) Cleavage site detection. The cleavage site (if any) of a query sequence is determined by a signal-based method.
2) Pre-sequence selection. The pre-sequence of the query is obtained by selecting from the N-terminal up to the cleavage site.
3) Pairwise alignment. The pre-sequence is aligned with each of the training pre-sequences to form an N-dim vector, which is fed to a one-vs-rest SVM classifier for prediction.
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Fusion of Signal- and Homology-Based Methods
Signal-based Cleavage Site Predictor
N
truncate
Homology-based Method
SubcellularLocation
CAmino Acid Sequence
…
Pre-sequence
Cleavage Site
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Perturbational Discriminant AnalysisSpectral Space:
Define the kernel matrix
K can be factorized via spectral decomposition into
Empirical Space
Nx )(e),( 1 xxK
Spectral Space