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hyginn 2017-10-22 23:38:32 -04:00
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BIN-ALI-Optimal_sequence_alignment.R
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@ -10,43 +10,57 @@
# #
# Versions: # Versions:
# 0.1 First code copied from 2016 material. # 0.1 First code copied from 2016 material.
# #
# TODO: # TODO:
# #
# #
# == DO NOT SIMPLY source() THIS FILE! ======================================= # == DO NOT SIMPLY source() THIS FILE! =======================================
#
# If there are portions you don't understand, use R's help system, Google for an # If there are portions you don't understand, use R's help system, Google for an
# answer, or ask your instructor. Don't continue if you don't understand what's # answer, or ask your instructor. Don't continue if you don't understand what's
# going on. That's not how it works ... # going on. That's not how it works ...
#
# ============================================================================== # ==============================================================================
# = 1 Biostrings Pairwise Alignment #TOC> ==========================================================================
#TOC>
#TOC> Section Title Line
#TOC> -------------------------------------------------------
#TOC> 1 Prepare 41
#TOC> 2 Biostrings Pairwise Alignment 49
#TOC> 2.1 Optimal global alignment 60
#TOC> 2.2 Optimal local alignment 123
#TOC> 3 APSES Domain annotation by alignment 147
#TOC> 4 Update your database script 228
#TOC>
#TOC> ==========================================================================
# Biostrings is one of the basic packages that the Bioconductor software
# landscape builds on. It stores sequences in "AAstring" objects and these are
# complex software structures that are designed to be able to handle
# genome-scale sequences. Biostrings functions - such as the alignment functions
# - expect their input to be Biostrings objects.
?AAString
AAString("ACDE")
s <- AAString("ACDE")
str(s)
# See: it's complicated. This is an "S4" object. Bioconductor uses these objects
# almost exclusively, but we will not be poking around in their internals. Just
# this: how do we get the sequence back out of an AAString object? The help page
# for XString - the parent "class" of AAStrings - mentions the alternatives:
as.character(s) # the base R version
toString(s) # using the Biostrings function toString()
# While we need to remember to convert our sequences from the character vectors # = 1 Prepare =============================================================
# that we store in our database, to AAStrings that we can align, the alignment
# itself is really straightforward. The pairwiseAlignment() function was written # You need to recreate the protein database that you have constructed in the
# to behave exactly like the functions you encountered on the EMBOSS server. # BIN-Storing_data unit.
source("makeProteinDB.R")
# = 2 Biostrings Pairwise Alignment =======================================
if (!require(Biostrings, quietly=TRUE)) {
source("https://bioconductor.org/biocLite.R")
biocLite("Biostrings")
library(Biostrings)
}
# Biostrings stores sequences in "XString" objects. Once we have onverted our
# traget sequences to AAString objects, the alignment itself is straightforward.
# == 2.1 Optimal global alignment ==========================================
# The pairwiseAlignment() function was written to behave
# exactly like the functions you encountered on the EMBOSS server.
# First: make AAString objects ... # First: make AAString objects ...
sel <- myDB$protein$name == "MBP1_SACCE" sel <- myDB$protein$name == "MBP1_SACCE"
@ -56,9 +70,8 @@ sel <- myDB$protein$name == paste("MBP1_", biCode(MYSPE), sep = "")
aaMBP1_MYSPE <- AAString(myDB$protein$sequence[sel]) aaMBP1_MYSPE <- AAString(myDB$protein$sequence[sel])
?pairwiseAlignment ?pairwiseAlignment
# ... and align. # ... and align.
# Global optimal alignment with end-gap penalties is default. (like EMBOSS needle) # Global optimal alignment with end-gap penalties is default.
ali1 <- pairwiseAlignment( ali1 <- pairwiseAlignment(
aaMBP1_SACCE, aaMBP1_SACCE,
aaMBP1_MYSPE, aaMBP1_MYSPE,
@ -66,7 +79,7 @@ ali1 <- pairwiseAlignment(
gapOpening = 10, gapOpening = 10,
gapExtension = 0.5) gapExtension = 0.5)
str(ali1) # Did you think the AAString object was complicated ? str(ali1) # ... it's complicated
# This is a Biostrings alignment object. But we can use Biostrings functions to # This is a Biostrings alignment object. But we can use Biostrings functions to
# tame it: # tame it:
@ -107,6 +120,8 @@ percentID <- function(al) {
percentID(ali1) percentID(ali1)
# == 2.2 Optimal local alignment ===========================================
# Compare with local optimal alignment (like EMBOSS Water) # Compare with local optimal alignment (like EMBOSS Water)
ali2 <- pairwiseAlignment( ali2 <- pairwiseAlignment(
aaMBP1_SACCE, aaMBP1_SACCE,
@ -117,8 +132,8 @@ ali2 <- pairwiseAlignment(
gapExtension = 10) gapExtension = 10)
writePairwiseAlignments(ali2) # This has probably only aligned the N-terminal writePairwiseAlignments(ali2) # This has probably only aligned the N-terminal
# DNA binding domain - but that one has quite # DNA binding domain - but that one has quite
# high sequence identity: # high sequence identity:
percentID(ali2) percentID(ali2)
# == TASK: == # == TASK: ==
@ -128,73 +143,49 @@ percentID(ali2)
# the gap penalties and see what happens: how does the number of indels change, # the gap penalties and see what happens: how does the number of indels change,
# how does the length of indels change... # how does the length of indels change...
# Fine. Please return to the Wiki to study BLAST alignment...
# = 3 APSES Domain annotation by alignment ================================
# ==============================================================================
# PART FOUR: APSES Domain annotation by alignment
# ==============================================================================
# In this section we define the MYSPE APSES sequence by performing a global, # In this section we define the MYSPE APSES sequence by performing a global,
# optimal sequence alignment of the yeast domain with the full length protein # optimal sequence alignment of the yeast APSES domain with the full length
# sequence of the protein that was the most similar to the yeast APSES domain. # protein sequence of the protein that was the most similar to the yeast APSES
# domain.
# #
# I have annotated the yeast APSES domain as a proteinAnnotation in the # I have annotated the yeast APSES domain as a feature in the
# database. To view the annotation, we can retrieve it via the proteinID and # database. To view the annotation, we can retrieve it via the proteinID and
# featureID. Here is the yeast protein ID: # featureID. Here is the yeast protein ID:
myDB$protein$ID[myDB$protein$name == "MBP1_SACCE"] (proID <- myDB$protein$ID[myDB$protein$name == "MBP1_SACCE"])
# ... assign it for convenience:
proID <- myDB$protein$ID[myDB$protein$name == "MBP1_SACCE"]
# ... and if you look at the feature table, you can identify the feature ID # ... and if you look at the feature table, you can identify the feature ID
myDB$feature[ , c("ID", "name", "description")] (ftrID <- myDB$feature$ID[myDB$feature$name == "APSES fold"])
myDB$feature$ID[myDB$feature$name == "APSES fold"]
# ... assign it for convenience: # ... and with the two annotations we can get the corresponding ID from the
ftrID <- myDB$feature$ID[myDB$feature$name == "APSES fold"]
# ... and with the two annotations we can pull the entry from the protein
# annotation table # annotation table
(fanID <- myDB$annotation$ID[myDB$annotation$proteinID == proID &
myDB$annotation$featureID == ftrID])
myDB$proteinAnnotation[myDB$proteinAnnotation$protein.ID == proID & myDB$proteinAnnotation[myDB$proteinAnnotation$protein.ID == proID &
myDB$proteinAnnotation$feature.ID == ftrID, ] myDB$proteinAnnotation$feature.ID == ftrID, ]
myDB$proteinAnnotation$ID[myDB$proteinAnnotation$protein.ID == proID &
myDB$proteinAnnotation$feature.ID == ftrID]
# ... assign it for convenience:
fanID <- myDB$proteinAnnotation$ID[myDB$proteinAnnotation$protein.ID == proID &
myDB$proteinAnnotation$feature.ID == ftrID]
# The annotation record contains the start and end coordinates which we can use # The annotation record contains the start and end coordinates which we can use
# to define the APSES domain sequence with a substr() expression. # to define the APSES domain sequence with a substr() expression.
substr(myDB$protein$sequence[myDB$protein$ID == proID],
myDB$proteinAnnotation$start[myDB$proteinAnnotation$ID == fanID], (start <- myDB$annotation$start[myDB$annotation$ID == fanID])
myDB$proteinAnnotation$end[myDB$proteinAnnotation$ID == fanID]) (end <- myDB$annotation$end[myDB$annotation$ID == fanID])
(apses <- substr(myDB$protein$sequence[myDB$protein$ID == proID],
start,
end))
# Lots of code. But don't get lost. Let's recapitulate what we have done: we # Lots of code. But don't get lost. Let's recapitulate what we have done: we
# have selected from the sequence column of the protein table the sequence whose # have selected from the sequence column of the protein table the sequence whose
# name is "MBP1_SACCE", and selected from the proteinAnnotation table the start # name is "MBP1_SACCE", and selected from the annotation table the start
# and end coordinates of the annotation that joins an "APSES fold" feature with # and end coordinates of the annotation that joins an "APSES fold" feature with
# the sequence, and used the start and end coordinates to extract a substring. # the sequence, and used the start and end coordinates to extract a substring.
# The expressions get lengthy, but it's not hard to wrap all of this into a
# function so that we only need to define name and feature.
dbGetFeatureSequence
dbGetFeatureSequence(myDB, "MBP1_SACCE", "APSES fold")
# Let's convert this to an AAstring and assign it: # Let's convert this to an AAstring and assign it:
aaMB1_SACCE_APSES <- AAString(dbGetFeatureSequence(myDB, aaMB1_SACCE_APSES <- AAString(apses)
"MBP1_SACCE",
"APSES fold"))
# To align, we need the MYSPE sequence. Here is it's definition again, just
# in case ...
sel <- myDB$protein$name == paste("MBP1_", biCode(MYSPE), sep = "")
aaMBP1_MYSPE <- AAString(myDB$protein$sequence[sel])
# Now let's align these two sequences of very different length without end-gap # Now let's align these two sequences of very different length without end-gap
# penalties using the "overlap" type. "overlap" turns the # penalties using the "overlap" type. "overlap" turns the
@ -233,57 +224,57 @@ aliApses@subject@range@start
# ... and end is: # ... and end is:
aliApses@subject@range@start + aliApses@subject@range@width - 1 aliApses@subject@range@start + aliApses@subject@range@width - 1
# Since we have this section defined now, we can create a feature annotation
# right away and store it in myDB. Copy the code-template below to your # = 4 Update your database script =========================================
# myCode.R file, edit it to replace the placeholder items with your data:
# Since we have this feature defined now, we can create a feature annotation
# right away and store it in myDB. Follow the following steps carefully:
# #
# - The <PROTEIN ID> is to be replaced with the ID of MBP1_MYSPE
# - The <FEATURE ID> is to be replaced with the ID of "APSES fold"
# - <START> and <END> are to be replaced with the coordinates you got above
# #
# Then execute the code and continue below the code template. If you make an # - Make a copy of the file "./data/refAnnotations.json" in your project
# error, there are instructions on how to recover, below. # directory and give it a new name that corresponds to MYSPE - e.g. if
# MYSPE is called "Crptycoccus neoformans", your file should be called
# "CRYNEAnnotations.json"; in that case "MBP1_CRYNE" would also be the
# "name" of your protein. Open the file in the RStudio editor and delete
# all annotations but one for an "APSES fold". Edit that annotation to
# correspond to the your MBP1_MYSPE protein and enter the start end end
# coordinates you have just discovered for the APSES domain in your
# sequence. Save your file.
# #
# ===== begin code template: add a proteinAnnotation to the database ===== # - Validate your file online at https://jsonlint.com/
#
# - Next, you need to update your "makeProteinDB.R" script to load the
# annotation when you recreate the database. Open the script in the
# RStudio ediotr, and add the following command at the end:
#
# myDB <- dbAddAnnotation(myDB, fromJSON("<MYSPE>Annotations.json"))
#
# - save the file and source() it:
#
# source("makeProteinDB.R")
#
# This should run without errors or warnings. If it doesn't work and you
# can't figure out quickly what's happeneing, ask on the mailing list for
# help.
#
# - Confirm
# The following commands should retrieve the correct start and end
# coordinates for the MBP1_MYSPE APSES domain:
# == edit all placeholder items! sel <- myDB$protein$name == paste("MBP1_", biCode(MYSPE), sep = "")
myProteinID <- "<PROTEIN ID>" aaMBP1_MYSPE <- AAString(myDB$protein$sequence[sel])
myFeatureID <- "<FEATURE ID>"
myStart <- <START>
myEnd <- <END>
# == create the proteinAnnotation entry
panRow <- data.frame(ID = dbAutoincrement(myDB$proteinAnnotation$ID),
protein.ID = myProteinID,
feature.ID = myFeatureID,
start = myStart,
end = myEnd,
stringsAsFactors = FALSE)
myDB$proteinAnnotation <- rbind(myDB$proteinAnnotation, panRow)
# == check that this was successful and has the right data
myDB$proteinAnnotation[nrow(myDB$proteinAnnotation), ]
# ===== end code template ===========================================
# ... continue here.
# I expect that a correct result would look something like
# ID protein.ID feature.ID start end
# 63 my_fan_1 my_pro_1 ref_ftr_1 6 104
# If you made a mistake, simply overwrite the current version of myDB by loading
# your saved, good version: load("myDB.01.RData") and correct your mistake
# If this is correct, save it
save(myDB, file = "myDB.02.RData") # Note that it gets a new version number!
# Done with this part. Copy the sequence of the APSES domain of MBP1_MYSPE - you
# need it for the reverse BLAST search, and return to the course Wiki.
(proID <- myDB$protein$ID[myDB$protein$name == "MBP1_<MYSSPE>"]) # <<< EDIT
# = 1 Tasks (ftrID <- myDB$feature$ID[myDB$feature$name == "APSES fold"])
(fanID <- myDB$annotation$ID[myDB$annotation$proteinID == proID &
myDB$annotation$featureID == ftrID])
(start <- myDB$annotation$start[myDB$annotation$ID == fanID])
(end <- myDB$annotation$end[myDB$annotation$ID == fanID])
(apses <- substr(myDB$protein$sequence[myDB$protein$ID == proID],
start,
end))