Question: Protein quantification by MSnbase package
0
gravatar for fsgolestan
3 months ago by
fsgolestan10
fsgolestan10 wrote:

Hello, I received a dataset (from proteome discoverer) containing the below information for both protein and peptide levels:

Protein level: Accession
Description Exp. q-value
Sum PEP Score
Coverage
Peptides
PSMs
Unique Peptides AAs MW [kDa]
calc. pI
Score Sequest HT
Peptides Sequest HT


Peptide level:

Annotated Sequence
Master Protein Accessions Missed Cleavages
Charge
DeltaScore
DeltaCn Rank
Search Engine Rank
m/z [Da]
MH+ [Da]
DeltaM [ppm]
Deltam/z [Da]
Activation Type MS Order
Isolation Interference [%]
Ion Inject Time [ms]
RT [min]
First Scan
Spectrum File
Ions Matched
XCorr
Percolator q-Value
Percolator PEP

However, there is no quantitative values of proteins that enables me to do deferentially expression analysis, and …. May I ask whether it is possible to perform protein quantitation from this dataset by MSnbase package? If the quantification of protein values is possible from my dataset, may I ask which of the above features can be used for calculation of protein values?

I would highly appreciate great help in this regard.

msnbase rforproteomics • 130 views
ADD COMMENTlink modified 12 weeks ago • written 3 months ago by fsgolestan10

Yes, you could do quantitation in MSnbase, depending on your MS quantitation methodology employed. However, from you mail above, it doesn't seem that you have the raw data anyway. Also, it is unclear what quantitation methodology was used - it could be spectral counting, or may be the quantitation values are stored in the Sequest variables? I would be useful to clarify what kind of quantitative data you have received.

ADD REPLYlink written 3 months ago by Laurent Gatto1.1k
Answer: Protein quantification by MSnbase package
0
gravatar for fsgolestan
12 weeks ago by
fsgolestan10
fsgolestan10 wrote:

Thanks for your reply. This dataset is only an identification dataset, and I asked the person in charge of running MS about protein quantitation values. He said this experiment is not designed for protein quantitation. I do not know why. Maybe because it is an immuno-precipitation experiment. The only information available for identified proteins and peptides in this dataset is from my mail above. I would like to know if by using these information, can I quantify proteins (semi-quantify) for example through spectral counting? or doing any quantitation methods is impossible for this dataset?

Thanks a lot for your help.

ADD COMMENTlink written 12 weeks ago by fsgolestan10

Yes, if you have identification only, you could do spectral counting - see also MSnID (requiring raw data though). In MSnbase, the quantify method support spectral counting.

ADD REPLYlink written 12 weeks ago by Laurent Gatto1.1k

Thank you very much. It is very good that I can do quantification from this identification dataset, while I do not have raw data. May I also know for quantification through spectral counting using MSnbase, which of the features (from my above list) will be used to do spectral counting and quantification? I would like to know which of these features in my dataset are the important features that MSnbase will use to obtain spectral counts and subsequently quantitation values?

Also, to do this task, may I know if, by using MSnID package, I should firstly read my MS identification data (csv) into R (converting csv to MSnID object), and then, convert MSnID object into mzIdentML (mzid) to be able to start for quantification analysis by MSnbase?

Many thanks for your great help.

ADD REPLYlink written 12 weeks ago by fsgolestan10

I think you will have to start from mzID files to use MSnID.

As for spectral counting using MSnbase, I would that you would want to count the reliable PSMs (based on their q-value) that have been associated to a protein group. So the fcol argument should probably be Accession.

ADD REPLYlink written 12 weeks ago by Laurent Gatto1.1k

Thanks a lot for your useful help.

ADD REPLYlink written 12 weeks ago by fsgolestan10
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