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peptide setting on enzyme characteristics

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peptide setting on enzyme characteristics lourdes thevanayagam  2018-06-07 15:41
 

Hello Brendan,

Thanks for the training on skyline at Buck Inst,
After the training I started working on protein quantification using peptides cleaved by different enzymes and it is working things getting better.

I have a question on peptide settings creating my own enzymatic digestion and on “edit current enzyme” I would like to have a clear understanding of choosing

  1.  Cleave C-terminal to

  2.  Cleave N-terminal to

I know where the enzyme cut the peptide

Example -----SGRSD------ the enzyme I chose cut RS, so in that case could you please advise me for the cleave C- terminal to and cleave N-terminal to.

Hope you understand my question.

Thanks for your help.

Sincerely
Lourdes
 
 
Brendan MacLean responded:  2018-06-07 16:23
Hi Lourdes,
I used the definition as follows:

Type = C-terminal
Cleave C-terminal to = R
Unless followed by = ACDEFGHIKLMNOPQRTUVWY

Equivalent to:

Type = N-terminal
Cleave N-terminal to = S
Unless followed by = ACDEFGHIKLMNOPQSTUVWY

These seemed to work for me in adding an arbitrarily chosen protein sequence as follows:

>IPI00000044 IPI:IPI00000044.1|SWISS-PROT:P01127|TREMBL:P78450;Q6FHE7|REFSEQ_NP:NP_002599|ENSEMBL:ENSP00000330382|H-INV:HIT000040889|VEGA:OTTHUMP00000028573 Tax_Id=9606 Platelet-derived growth factor B chain precursor
MNRCWALFLSLCCYLRLVSAEGDPIPEELYEMLSDHSIRSFDDLQRLLHGDPGEEDGAEL
DLNMTRSHSGGELESLARGRRSLGSLTIAEPAMIAECKTRTEVFEISRRLIDRTNANFLV
WPPCVEVQRCSGCCNNRNVQCRPTQVQLRPVQVRKIEIVRKKPIFKKATVTLEDHLACKC
ETVAAARPVTRSPGGSQEQRAKTPQTRVTIRTVRVRRPPKGKHRKFKHTHDKTALKETLG
A*
    R.SHSGGELESLARGRR.S [66, 80]

Is this what you had in mind? Hope this helps to explain the intent of the fields in the Edit Enzyme form.

Thanks for posting to the Skyline support board.

--Brendan