Automated resonance assignment with FLYA (Gothenburg 2014) - Revision history

Guentert: /* Fully automated structure calculation */

2014-10-14T08:00:55Z

Fully automated structure calculation

← Older revision		Revision as of 10:00, 14 October 2014
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	Automated resonance assignment, automated NOE restraint assignment, and the structure calculation by torsion angle dynamics can be combined by running the 'flya' command in 'CALC.cya' with the additional parameter 'stage=1' (which was commented out so far):		Automated resonance assignment, automated NOE restraint assignment, and the structure calculation by torsion angle dynamics can be combined by running the 'flya' command in 'CALC.cya' with the additional parameter 'stage=1' (which was commented out so far):

	flya ~~refprot~~=ref.prot ~~noesy~~=$~~noesy scalar~~=$~~scalar runs=5~~ stage=1		flya runs=10 shiftreference=ref.prot structurepeaks=$structurepeaks assignpeaks=$assignpeaks stage=1

	The 'flya.prot' file from the automated resonance assignment will be used together with the (unassigned) NOESY peak lists to assign the NOESY peaks and to generate distance restraints in order to compute the three-dimensional structure of the protein.		The 'flya.prot' file from the automated resonance assignment will be used together with the (unassigned) NOESY peak lists to assign the NOESY peaks and to generate distance restraints in order to compute the three-dimensional structure of the protein.

Guentert: /* Run the FLYA calculation */

2014-10-14T07:33:25Z

Run the FLYA calculation

← Older revision		Revision as of 09:33, 14 October 2014
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	To run the FLYA calculation, you start CYANA and execute the 'CALC.cya' script:		To run the FLYA calculation, you start CYANA and execute the 'CALC.cya' script:

	cyana "nproc=5; CALC"		cyana "nproc=2; CALC"

	By specifying the 'nproc=5' command, the 5 independent runs of the algorithm will be performed in parallel. On a computer with multiple processors this will speed up the calculation, which is expected to take a few minutes.		By specifying the 'nproc=2' command, the independent runs of the algorithm will be performed in parallel. On a computer with multiple processors this will speed up the calculation, which is expected to take a few minutes.

	== FLYA output files ==		== FLYA output files ==

Guentert: /* Experiment definitions in the CYANA library */

2014-10-14T07:32:05Z

Experiment definitions in the CYANA library

← Older revision		Revision as of 09:32, 14 October 2014
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	SPECTRUM HNCA HN N C		SPECTRUM HNCA HN N C
	0.980 HN:H_AMI N:N_AM* ~~C_A* C_BYL~~ C:C_ALI		0.980 HN:H_AMI N:N_AM* C:C_ALI C_BYL
	0.800 HN:H_AMI N:N_AMI C_BYL ~~C_ALI N_AMI~~ C:C_ALI		0.800 HN:H_AMI N:N_AMI (C_ALI) C_BYL C:C_ALI

	The first line corresponds to the '#SPECTRUM' line in the peak list. It specifies the experiment name and a label for the atoms that are detected in each dimension of the spectrum. The number of labels defines the dimensionality of the experiment (3 in case of HNCA).		The first line corresponds to the '#SPECTRUM' line in the peak list. It specifies the experiment name and a label for the atoms that are detected in each dimension of the spectrum. The number of labels defines the dimensionality of the experiment (3 in case of HNCA).

Guentert: /* CYANA setup for the NMR course in Gothenburg (15 October 2014) */

2014-10-14T07:29:30Z

CYANA setup for the NMR course in Gothenburg (15 October 2014)

← Older revision		Revision as of 09:29, 14 October 2014
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	Please follow the following steps carefully (exact Linux commands are given below; you may copy them to a terminal):		Please follow the following steps carefully (exact Linux commands are given below; you may copy them to a terminal):

	# ~~From~~ your home directory, ~~change to the~~ directory '~~guentert~~'.		# In your home directory, make a directory 'cyana' and change into it.
	~~# Execute the setup script~~.
	# Download the [[Media:flyadata.tgz\|input data for the practical]].		# Download the [[Media:flyadata.tgz\|input data for the practical]].
	# Unpack the input data for the practical.		# Unpack the input data for the practical.
	# ~~Move~~ it to different directory, '~~flyaembo1~~' (~~such that you later unpack~~ the original data ~~again~~ to run another calculation)		# Copy it to different directory, 'flya01' (to keep the original data in 'flyadata' to run another calculation later)
	# Change into the subdirectory '~~flyaembo1~~' where the input data for the practical are.		# Change into the subdirectory 'flya01' where the input data for the practical are.
	# Test whether CYANA can be started by typing its name, 'cyana'.		# Test whether CYANA can be started by typing its name, 'cyana'.
	# Exit from CYANA by typing 'q' or 'quit'.		# Exit from CYANA by typing 'q' or 'quit'.

	cd ~		cd ~
	wget 'http://www.cyana.org/wiki/images/3/31/~~Flyaembo~~.tgz'		mkdir cyana
			cd cyana
			wget 'http://www.cyana.org/wiki/images/4/46/Flyadata.tgz'
	tar zxf Flyadata.tgz		tar zxf Flyadata.tgz
	cp -a flyadata flya01		cp -a flyadata flya01
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	Demo license valid for specific sequences until 2014-12-31.		Demo license valid for specific sequences until 2014-12-31.
	Library file "/Users/guentert/~~Documents/Meetings/1410NMRCourseGothenburg/Practical~~/cyana-3.97/lib/cyana.lib" read, 38 residue types.		Library file "/Users/guentert/bin/cyana-3.97/lib/cyana.lib" read, 38 residue types.
	Sequence file "demo.seq" read, 114 residues.		Sequence file "demo.seq" read, 114 residues.
	cyana> q		cyana> q

Guentert: /* CYANA setup for the NMR course in Gothenburg (15 October 2014) */

2014-10-14T07:25:13Z

CYANA setup for the NMR course in Gothenburg (15 October 2014)

← Older revision		Revision as of 09:25, 14 October 2014
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	# From your home directory, change to the directory 'guentert'.		# From your home directory, change to the directory 'guentert'.
	# Execute the setup script.		# Execute the setup script.
	# Download the [[Media:~~flyaembo~~.tgz\|input data for the practical]].		# Download the [[Media:flyadata.tgz\|input data for the practical]].
	# Unpack the input data for the practical.		# Unpack the input data for the practical.
	# Move it to different directory, 'flyaembo1' (such that you later unpack the original data again to run another calculation)		# Move it to different directory, 'flyaembo1' (such that you later unpack the original data again to run another calculation)
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	# Exit from CYANA by typing 'q' or 'quit'.		# Exit from CYANA by typing 'q' or 'quit'.

	cd ~~~/guentert~~		cd ~
	~~source setup-guentert~~
	wget 'http://www.cyana.org/wiki/images/3/31/Flyaembo.tgz'		wget 'http://www.cyana.org/wiki/images/3/31/Flyaembo.tgz'
	tar zxf ~~Flyaembo~~.tgz		tar zxf Flyadata.tgz
	~~mv flyaembo flyaembo1~~		cp -a flyadata flya01
	cd ~~flyaembo1~~		cd flya01
	cyana		cyana
	___________________________________________________________________		___________________________________________________________________

Guentert at 07:14, 14 October 2014

2014-10-14T07:14:31Z

← Older revision		Revision as of 09:14, 14 October 2014
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	== Run the FLYA calculation ==		== Run the FLYA calculation ==

	To run the FLYA calculation, you start CYANA and execute the '~~RUNFLYA~~.cya' script:		To run the FLYA calculation, you start CYANA and execute the 'CALC.cya' script:

	cyana "nproc=5; ~~RUNFLYA~~"		cyana "nproc=5; CALC"

	By specifying the 'nproc=5' command, the 5 independent runs of the algorithm will be performed in parallel. On a computer with multiple processors this will speed up the calculation, which is expected to take a few minutes.		By specifying the 'nproc=5' command, the 5 independent runs of the algorithm will be performed in parallel. On a computer with multiple processors this will speed up the calculation, which is expected to take a few minutes.
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	=== The flya.txt file ===		=== The flya.txt file ===

	This output file starts with overall assignment statistics for each group of atoms as defined by '~~analyze_assign_group~~:=...' in ~~RUNFLYA~~.cya':		This output file starts with overall assignment statistics for each group of atoms as defined by 'analyzeassign_group:=...' in CALC.cya':

	____________________________________________________________		____________________________________________________________
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	shiftassign_fix:=fix.prot		shiftassign_fix:=fix.prot
	noesy:=N15NOESY,C13NOESY		noesy:=N15NOESY,C13NOESY
	~~scalar~~:=C13H1,N15H1,HCCH24,HCCH7,HBHACONH,C_CO_NH,HC_CO_NH		assignpeaks:=C13H1,N15H1,HCCH24,HCCH7,HBHACONH,C_CO_NH,HC_CO_NH

	== Fully automated structure calculation ==		== Fully automated structure calculation ==

	Automated resonance assignment, automated NOE restraint assignment, and the structure calculation by torsion angle dynamics can be combined by running the 'flya' command in '~~RUNFLYA~~.cya' with the additional parameter 'stage=1' (which was commented out so far):		Automated resonance assignment, automated NOE restraint assignment, and the structure calculation by torsion angle dynamics can be combined by running the 'flya' command in 'CALC.cya' with the additional parameter 'stage=1' (which was commented out so far):

	flya refprot=ref.prot noesy=$noesy scalar=$scalar runs=5 stage=1		flya refprot=ref.prot noesy=$noesy scalar=$scalar runs=5 stage=1
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	The 'flya.prot' file from the automated resonance assignment will be used together with the (unassigned) NOESY peak lists to assign the NOESY peaks and to generate distance restraints in order to compute the three-dimensional structure of the protein.		The 'flya.prot' file from the automated resonance assignment will be used together with the (unassigned) NOESY peak lists to assign the NOESY peaks and to generate distance restraints in order to compute the three-dimensional structure of the protein.

	To speed up the calculation, you can set in '~~RUNFLYA~~.cya' (above the 'flya' command):		To speed up the calculation, you can set in 'CALC.cya' (above the 'flya' command):

	structures:=25,5		structures:=25,5

Guentert: /* Using input chemical shifts: shift predictions or partial assignments (optional) */

2014-10-13T21:31:00Z

Using input chemical shifts: shift predictions or partial assignments (optional)

← Older revision		Revision as of 23:31, 13 October 2014
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	These shifts will only be used for comparison (e.g. in flya.tab, flya.txt, flya.pdf):		These shifts will only be used for comparison (e.g. in flya.tab, flya.txt, flya.pdf):

	~~run_assign_reference~~:=ref.prot		shiftassign_reference:=ref.prot

	Shifts and standard deviations in the file 'predicted.prot' (not provided in this practical) will replace the general statistics from cyana.lib (CSTABLE):		Shifts and standard deviations in the file 'predicted.prot' (not provided in this practical) will replace the general statistics from cyana.lib (CSTABLE):

	~~run_assign_statistics~~:=predicted.prot		shiftassign_statistics:=predicted.prot

	Shifts in the file 'fix.prot' will be fixed to the input values		Shifts in the file 'fix.prot' will be fixed to the input values

	~~run_assign_fix~~:=fix.prot		shiftassign_fix:=fix.prot

	The latter approach can for instance be used to perform sidechain assignment when the backbone assignment is already known.		The latter approach can for instance be used to perform sidechain assignment when the backbone assignment is already known.
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	q		q

	The file 'fix.prot' will contain the reference chemical shifts only for the backbone (and CB) atoms H, N, CA, CB, C'. Now you can repeat the assignment calculation by inserting the '~~run_assign_fix~~:=fix.prot' statement in '~~RUNFLYA~~.cya' and choosing only the input peak lists that are relevant for sidechain assignment:		The file 'fix.prot' will contain the reference chemical shifts only for the backbone (and CB) atoms H, N, CA, CB, C'. Now you can repeat the assignment calculation by inserting the 'shiftassign_fix:=fix.prot' statement in 'CALC.cya' and choosing only the input peak lists that are relevant for sidechain assignment:

	~~run_assign_fix~~:=fix.prot		shiftassign_fix:=fix.prot
	noesy:=N15NOESY,C13NOESY		noesy:=N15NOESY,C13NOESY
	scalar:=C13H1,N15H1,HCCH24,HCCH7,HBHACONH,C_CO_NH,HC_CO_NH		scalar:=C13H1,N15H1,HCCH24,HCCH7,HBHACONH,C_CO_NH,HC_CO_NH

Guentert: /* FLYA output files */

2014-10-13T21:29:16Z

FLYA output files

Show changes

Guentert: /* FLYA execution script */

2014-10-13T21:22:22Z

FLYA execution script

← Older revision		Revision as of 23:22, 13 October 2014
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	structurepeaks:=N15NOESY,C13NOESY		structurepeaks:=N15NOESY,C13NOESY
	#assignpeaks:=N15NOESY,C13NOESY,C13HSQC,N15HSQC,HCCHTOCSY,HCCHCOSY,HNCA,HNcaCO,HNCO,HNcoCA,CBCANH,CBCAcoNH,HBHAcoNH,CcoNH,HCcoNH		assignpeaks:=N15NOESY,C13NOESY,C13HSQC,N15HSQC,HCCHTOCSY,HCCHCOSY,HNCA,HNcaCO,HNCO,HNcoCA,CBCANH,CBCAcoNH,HBHAcoNH,CcoNH,HCcoNH
	~~assignpeaks:=N15HSQC,HNCA,HNcaCO,HNCO,HNcoCA,CBCANH,CBCAcoNH~~

	Experiments used for automated NOESY assignment and structure calculation are given in the 'structurepeaks:=...' statement (not relevant for the present practical); those for automated chemical shift assignment in the 'assignpeaks:=...' statement. The peak list names are separated by commas (without blanks!).		Experiments used for automated NOESY assignment and structure calculation are given in the 'structurepeaks:=...' statement (not relevant for the present practical); those for automated chemical shift assignment in the 'assignpeaks:=...' statement. The peak list names are separated by commas (without blanks!).
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	The commands above will use all available peak lists. You can choose any subset of them by modifying the 'noesy:=...' or 'scalar:=...' statements.		The commands above will use all available peak lists. You can choose any subset of them by modifying the 'noesy:=...' or 'scalar:=...' statements.

	'''Try it!''' For instance, to make only backbone assignment using the triple resonance backbone assignment spectra, you can set		'''Try it!''' For instance, to make only backbone assignment using the <sup>15</sup>N-HSQC and triple resonance backbone assignment spectra, you can set

	~~noesy:=~~		assignpeaks:=N15HSQC,HNCA,HNcaCO,HNCO,HNcoCA,CBCANH,CBCAcoNH
	~~scalar~~:=HNCA,~~HN_CA_CO~~,HNCO,~~HN_CO_CA~~,CBCANH,~~CBCACONH,HBHACONH~~

	These are followed by tolerances for chemical shift matching:		These are followed by tolerances for chemical shift matching:
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	These define		These define

	* The population size for the genetic algorithm, i.e. how many assignments form one generation (20; chosen smaller than in normal production runs in order to speed up the calculation)		* The population size for the genetic algorithm, i.e. how many assignments form one generation (25; chosen smaller than in normal production runs in order to speed up the calculation)
	* The maximal number of iterations during local optimization (15000).		* The maximal number of iterations during local optimization (15000).
	* ~~A file with reference chemical shifts (ref.prot). These will not be used by~~ the ~~algorithm but only for a subsequent analysis of its results~~.		* An option to choose the "quick" optimization schedule.
	* Groups of atoms for which assignment statistics will be calculated and reported in the 'flya.txt' output file.		* Groups of atoms for which assignment statistics will be calculated and reported in the 'flya.txt' output file.
	* The seed value for the random number generator. Any positive integer number can be used.		* The seed value for the random number generator. Any positive integer number can be used.
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	Finally, there is the command to start the FLYA algorithm:		Finally, there is the command to start the FLYA algorithm:

	flya ~~refprot~~=ref.prot ~~noesy~~=$~~noesy scalar~~=$~~scalar runs=5 #stage=1~~		flya runs=10 shiftreference=ref.prot structurepeaks=$structurepeaks assignpeaks=$assignpeaks

	Here, the given parameters of the 'flya' command specify that		Here, the given parameters of the 'flya' command specify that

	* The results will be compared with the reference chemical shifts in the file 'ref.prot' (which have been determined independently by conventional methods).		* The number of independent runs of the algorithm, from which the consolidated shift will be calculated (chosen smaller than in normal production runs in order to speed up the calculation).
			* The results will be compared with the reference chemical shifts in the file 'ref.prot' (which have been determined independently by conventional methods). The reference chemical shifts will not be used by the algorithm but only for a subsequent analysis of its results.
	* The input peak lists that will be used (as defined above).		* The input peak lists that will be used (as defined above).
	* The number of independent runs of the algorithm, from which the consolidated shift will be calculated (chosen smaller than in normal production runs in order to speed up the calculation).
	<!--* Whether or not the resonance assignment and the NOESY peak lists will be used for a structure calculation-->		<!--* Whether or not the resonance assignment and the NOESY peak lists will be used for a structure calculation-->

Guentert at 21:16, 13 October 2014

2014-10-13T21:16:58Z

← Older revision		Revision as of 23:16, 13 October 2014
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	'''Hint:''' The formats of other CYANA files are described in the [[CYANA 3.0 Reference Manual]].		'''Hint:''' The formats of other CYANA files are described in the [[CYANA 3.0 Reference Manual]].

			<!--
	== FLYA initialization script ==		== FLYA initialization script ==

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	The command 'cyanalib' reads the standard CYANA library. The second command reads the protein sequence.		The command 'cyanalib' reads the standard CYANA library. The second command reads the protein sequence.
			-->
	== Experiment definitions in the CYANA library ==		== Experiment definitions in the CYANA library ==