PWM scores oscillating at highly regular intervals after ATG start codon

Question

[ahtmatrix]

I have an R script that takes in a fasta containing all coding sequences in human RNA. I then extracted 9 bases before the start codon of each CDS and the 1 base after the start codon. Here is an example sequence: "ATCGTAGCT ATG A".

I then used Biostrings in R to construct a PWM as shown below.

#make a pwm of length 13
human = readDNAStringSet('extracted_rna.fasta)
human.kozak = DNAStringSet(human)
human.pwm    = PWM(human.kozak, type = 'log2probratio')

I then used the human mRNA PWM to score the first 99 positions of a fasta of viral coding sequences below.

subject.fasta = readDNAStringSet('viral.fasta')

pwm.score.dataframe = NULL

#loop through length of subject DNAStringSet which should be num of sequences in fasta
for (i in 1:length(subject.fasta)) {

  #using the human pwm score using a sliding window of 13 as defined by the PWM until position 99 in subject seq
  scores <- PWMscoreStartingAt(human.pwm, subject.fasta[[i]], starting.at = 1:99)

  pwm.score.dataframe = cbind(pwm.score.dataframe, scores)
}

write.table(pwm.score.dataframe, file = "humanPWM_viral.csv")

When plotted in Microsoft Excel, I averaged the score of each position and graphed the position vs score below. Regardless of the subset of viral sequences I used, after the ATG spike the PWM scores oscillate with a wavelength of 3 positions. What is causing this behavior? You'd expect that after the the ATG start codon there would be enough random noise to cancel out fluctuations in the PWM score.

[Santosh Anand]

The result looks really strange to me. What were your reasons to do this analysis and what were you expecting to see?

[ddiez]

You say:

I then used the human mRNA PWM

But in your code I see:

#using the human pwm score using a sliding window of 13 as defined by the PWM until position 99 in subject seq
  scores <- PWMscoreStartingAt(virus.pwm, subject.fasta[[i]], starting.at = 1:99)

It seems instead of using human.pwm you are using virus.pwm. Could that be the source of your problems?

[ahtmatrix]

Ah thats a typo I reran it with the corrections and it gives the same result