ProDBWiki/WebDocumentation/PeptideMassFingerprintWithEmowse - Revision history

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2011-10-26T06:18:15Z

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imported>JomunaChoudhuri at 08:48, 24 August 2005

2005-08-24T08:48:49Z

← Older revision		Revision as of 10:48, 24 August 2005
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	Now you can specify some details for your search. You can title the search if you like. You can choose any taxonomy but it will not affect the search with '''''emowse'''''. The selection of the searchengine '''''emowse''''' and the database(s) to search against are mandatory. If you know the whole sequence molecular weight of your protein you can declare the weight in the field ''Protein Mass''. The unit is dalton. You can also select the enzyme or reagent used for the cleavage of the protein. If you are unsure which parameters you should specify in addition, do not do anything more. Otherwise you can specify the allowed whole seqence weight variability (Weight Variability), the error allowed for mass accuracy of experimental mass determination (Error Tolerance) and the weighting given to partially-cleaved peptide fragments. See more details in the [[#parameters\|Use of Parameters]] section.		Now you can specify some details for your search. You can title the search if you like. You can choose any taxonomy but it will not affect the search with '''''emowse'''''. The selection of the searchengine '''''emowse''''' and the database(s) to search against are mandatory. If you know the whole sequence molecular weight of your protein you can declare the weight in the field ''Protein Mass''. The unit is dalton. You can also select the enzyme or reagent used for the cleavage of the protein. If you are unsure which parameters you should specify in addition, do not do anything more. Otherwise you can specify the allowed whole seqence weight variability (Weight Variability), the error allowed for mass accuracy of experimental mass determination (Error Tolerance) and the weighting given to partially-cleaved peptide fragments. See more details in the [[#parameters\|Use of Parameters]] section.
	If you like to repeat a search with the same parameters, you have to choose the ''search again with old search parameters'' option at the bottom. All you have to do now is to click the ''Search'' button and '''''emowse''''' will do the rest. When the search is finished, you will see a new window with the number of hits. To see the results, click on ''Browse Results''.		If you like to repeat a search with the same parameters, you have to choose the ''search again with old search parameters'' option at the bottom. All you have to do now is to click the ''Search'' button and '''''emowse''''' will do the rest. When the search is finished, you will see a new window with the number of hits. To see the results, click on ''Browse Results''.

			<span id="parameters"></span>

	== Use of Parameters ==		== Use of Parameters ==

	~~<span id="parameters"></span>~~

	=== Databases ===		=== Databases ===
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	Here you can specifiy the whole sequence molecular weight (in dalton) for the given protein (if known). If no value is given, '''''emowse''''' will assign 0. This options allows you to limit the search to proteins of this molecular weight plus/minus a 'limit' (see [[#weight\|Weight Variability]]). Otherwise the whole database will be searched.		Here you can specifiy the whole sequence molecular weight (in dalton) for the given protein (if known). If no value is given, '''''emowse''''' will assign 0. This options allows you to limit the search to proteins of this molecular weight plus/minus a 'limit' (see [[#weight\|Weight Variability]]). Otherwise the whole database will be searched.

			<span id="weight"></span>

	=== Weight Variability ===		=== Weight Variability ===
	~~<span id="weight"></span>~~

	Here you can specify the allowed whole sequence weight variability. This option is meaningless without the ''Protein Mass'' parameter. If a molecular weight for the protein is specified, this option will limit the search to proteins with molecular weights +/- the given percentage. The default value is 25%. This value is suggested for initial searches, because discrimination is best when the filter percentage is narrow, but some molecular weight estimates (particularly from SDS gels) should be given considerable allowance for error. You can also specify a range of percentages for which searches are done. Example:		Here you can specify the allowed whole sequence weight variability. This option is meaningless without the ''Protein Mass'' parameter. If a molecular weight for the protein is specified, this option will limit the search to proteins with molecular weights +/- the given percentage. The default value is 25%. This value is suggested for initial searches, because discrimination is best when the filter percentage is narrow, but some molecular weight estimates (particularly from SDS gels) should be given considerable allowance for error. You can also specify a range of percentages for which searches are done. Example:

imported>AnnaKranz at 11:36, 3 June 2005

2005-06-03T11:36:18Z

← Older revision		Revision as of 13:36, 3 June 2005
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	* V8-phosph		* V8-phosph
	* Chymotrypsin		* Chymotrypsin
	* !CNBr		* CNBr

	You can only choose one enzyme/reagent for the searches. '''''Emowse''''' simulates the complete digests for the sequences in the databases using the following rules:		You can only choose one enzyme/reagent for the searches. '''''Emowse''''' simulates the complete digests for the sequences in the databases using the following rules:

Unknown user at 00:00, 1 January 1970

1970-01-01T00:00:00Z

New page

__NOTOC__
= Emowse - Protein Identification by mass spectrometry =

== Overview ==

For the unique and rapid identification of unknown sample proteins, a 'fingerprint' signature derived from peptide mass information is sufficient. By using the molecular weights of the peptides obtained by mass spectrometry, '''''emowse''''' searches a protein database for matches with the given data. The scoring algorithm tolerates experimental errors of a few Daltons. For each entry in the database to search, emowse derives whole sequence molecular weight and calculated peptide molecular weights for complete digests using a range of cleavage rules. If you like to look for your unknown sample using '''''emowse''''', the peptide molecular weights of the protein are sufficient. In addition you can specify a variety of other parameters such as cleavage enzyme, whole sequence molecular weight and error tolerance.

== How to use ==

If you like to search for your protein with '''''emowse''''' against a database, you have to select your Experiment and choose the link ''Mass Spectra Data'' on the left hand side of the web-page. There you have to select MS for scan type and choose your scans. Afterwards you get on to the search form by clicking on the ''Peptide Mass Fingerprint'' link.
Now you can specify some details for your search. You can title the search if you like. You can choose any taxonomy but it will not affect the search with '''''emowse'''''. The selection of the searchengine '''''emowse''''' and the database(s) to search against are mandatory. If you know the whole sequence molecular weight of your protein you can declare the weight in the field ''Protein Mass''. The unit is dalton. You can also select the enzyme or reagent used for the cleavage of the protein. If you are unsure which parameters you should specify in addition, do not do anything more. Otherwise you can specify the allowed whole seqence weight variability (Weight Variability), the error allowed for mass accuracy of experimental mass determination (Error Tolerance) and the weighting given to partially-cleaved peptide fragments. See more details in the [[#parameters|Use of Parameters]] section.
If you like to repeat a search with the same parameters, you have to choose the ''search again with old search parameters'' option at the bottom. All you have to do now is to click the ''Search'' button and '''''emowse''''' will do the rest. When the search is finished, you will see a new window with the number of hits. To see the results, click on ''Browse Results''.

== Use of Parameters ==

<span id="parameters"></span>

=== Databases ===

The offered databases are provided from the mascot server. You can select as many databases as you like. '''''Emowse''''' will search against each databases individually.

=== Enzyme/Reagent ===

'''''Emowse''''' provides the search with the following enzymes/reagents:
* Trypsin
* Lys-C
* Arg-C
* Asp-N
* V8-bicarb
* V8-phosph
* Chymotrypsin
* !CNBr

You can only choose one enzyme/reagent for the searches. '''''Emowse''''' simulates the complete digests for the sequences in the databases using the following rules:

<pre><nowiki>Reagent Cleavage rule
*********** ***************
Trypsin C-term to K/R
Lys-C C-term to K
Arg-C C-term to R
Asp-N N-term to D
V8-bicarb C-term to E
V8-phosph C-term to E/D
CNBr C-term to M
</nowiki></pre>

=== Protein Mass ===

Here you can specifiy the whole sequence molecular weight (in dalton) for the given protein (if known). If no value is given, '''''emowse''''' will assign 0. This options allows you to limit the search to proteins of this molecular weight plus/minus a 'limit' (see [[#weight|Weight Variability]]). Otherwise the whole database will be searched.

=== Weight Variability ===
<span id="weight"></span>

Here you can specify the allowed whole sequence weight variability. This option is meaningless without the ''Protein Mass'' parameter. If a molecular weight for the protein is specified, this option will limit the search to proteins with molecular weights +/- the given percentage. The default value is 25%. This value is suggested for initial searches, because discrimination is best when the filter percentage is narrow, but some molecular weight estimates (particularly from SDS gels) should be given considerable allowance for error. You can also specify a range of percentages for which searches are done. Example:

<pre><nowiki>If you specify 25% to 30% for Weight Variability, emowse will search each sample 6 times with different percentages for the allowed whole sequence molecular weight:
* 25%
* 26%
* 27%
* 28%
* 29%
* 30%
</nowiki></pre>

=== Error Tolerance ===

Here you can specify the error allowed for mass accuracy of experimental mass determination. If no value is specified, a default tolerance of 2 Daltons will be assumed. Be aware that descrimination will be improved by selecting a small error tolerance. You can also specify a range of error tolerances to search. '''''Emowse''''' will look for every tolerance given in steps of 0.1.

=== Partials Factor ===

Here you can specify the weighting given to parially-cleaved peptide fragments. The default value for ''Partials Factor'' is 0.4. This factor down-weights the score awarded to a partial fragment by the given amount.Example:

<pre><nowiki> A partials factor of 0.4 will reduce the score of partial fragments to 40% of the score of a complete peptide cleavage fragment of equal mass.
</nowiki></pre>

If the digestion is perfect (no partial fragments), you can optimize your search by specifiing a low partial factor. But if your data contains a significant proportion of partial cleavage fragments, you should set the partials factor to a higher value to improve discrimination.
As the parameters ''Weight Variabilit'' and ''Error Tolerance'' you can specify a range of partials factors as well. In steps of 0.1, '''''emowse''''' will search with every partials factor given in the range.

Author: Anna-Lena Kranz