Aringh at 15:47, 15 December 2009

2009-12-15T15:47:39Z

Alex: Created page with 'The minimum measurement time of (4,3)D GFT experiments can be reliably predicted from the S/N distribution of a 2D [15N, 1H] HSQC and the rotational correlation time tau_c. First…'

2009-11-30T17:26:06Z

Created page with 'The minimum measurement time of (4,3)D GFT experiments can be reliably predicted from the S/N distribution of a 2D [15N, 1H] HSQC and the rotational correlation time tau_c. First…'

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The minimum measurement time of (4,3)D GFT experiments can be reliably predicted from the S/N distribution of a 2D [15N, 1H] HSQC and the rotational correlation time tau_c. First, you need to generate an integrated peaklist.

==== '''Peak Integration in 2D [15N, 1H] HSQC''' ====

Having processed 2D (15N, 1H) HSQC as described in data process, one can do the peak picking and integration of 2D (15N, 1H) HSQC manualy or semi-automaticlly as following:

*Peak picking and integration by using program XEASY

##Use command <tt>ns</tt> to load the spectrum
##Use <tt>ls</tt> to load the corresponding sequence file (optional)
##Use <tt>in</tt> to automatic pick peaks with total peak number slightly more than expected by select adjust contour level; then manually remove side-chain amide peaks
##Use <tt>mn</tt> to measure noise level, and use <tt>tw</tt> to display the noise level value. Normally the value of 2.5 times of standard deviation ( ~250 if the noise level has been normalized ) is taken as noise level.
##Use <tt>ip</tt> to choose peak height (m) as integration mode, and use "ii" to integrate the whole spectrum. An [[NESG:XEASY|XEASY]] external program "PeakintI" can be used to obtain more accurate peak height values, which is discribed below.
##Use <tt>wp</tt> to save the peak list.

*More accurate peak height measurement by using program [[NESG:PeakintI|PeakintI]]. Type command: <tt>peakintI ../data/NHsqc6001 nhsqcsa_b.peaks 250 -i -t 2 2 0.1 </tt> where the <tt>nhsqcsa_b.peaks</tt> is the input peak list and the output file will be <tt>inhsqcsa_b.peaks</tt>.
*Combing atom name informaiton in the peak list for S/N distribution (optional). 
**by using [[NESG:UBNMR|UBNMR]], please check [[NESG:UBNMR|UBNMR]] macro
**by using script '''sim''', run macro '''comb_yang''' by typing <tt>sim comb_yang</tt>

==== '''S/N distribution analysis for 2D (15N, 1H) HSQC''' ====

The SN distribution of resonances in a NMR spectra can be fit to the Gaussian distribution:

[[Image:Sn_nhsqc.jpg]]

 <tt>f= a*exp(-0.5*((ln(SN_i)-ln(SN_0))/b)^2) </tt> where <tt>SN_0</tt> is the most populated S/N observed, <tt>f is the expected population at a certern SN value SN_i</tt>, <tt>a and b</tt> are contants.

The SN of NHSQC <tt>SN_0</tt> can be obtained as following:

#Calculate <tt>ln(SN)</tt> for each peak from the peak height and noise level, e.g. by using EXCEL
#Obtain SN distribution (<tt>population v.s. ls(SN)</tt>) by using Sigma-Plot
#By using Sigma-Plot, fitting SN distribution to Gaussian distribution and obtain <tt>SN_0</tt>, constant <tt>a and b</tt>.

==== '''Calculation of Measurement Time''' ====

The SN distribution of resonances in other NMR spectra can also be fit to the Gaussian distribution as in 2D (15N, 1H) NHSQC: <tt>f= a*exp(-0.5*((ln(SN_i)-ln(SN_0))/b)^2)</tt> where <tt>SN_0</tt> is the most populated SN observed, <tt>f</tt> is the expected population at a certain SN value <tt>SN_i</tt>, <tt>a</tt> and <tt>b</tt> are constants. Based on this equation, one can calculate the expected SN_0 for a required peak detection yield.

Assuming that a peak shall have at least SN value of 2 in order to be observed or detected, and the average b for (4,3) GFT experiments is 0.8; if 95% peak detection yield is required, the exptected SN_0 is: <tt>SN_0=exp(ln2+1.644*b)=7.4 </tt> 

[[Image:Sn2.jpg]]

NMR measurement time of (4,3) GFT HNNCABCA, (4,3)D GFT CABCAcoNHN and (4,3)D HABCABCONHN can be calculated from the following equation: <tt>T_43d= ((SN_43d* Tauc^2)/(SN_2d*A))^2 </tt> where <tt>T_43d</tt> is the required time for (4,3) GFT experiment, <tt>SN_43d</tt> is the expected SN value of (4,3)D GFT experiments, <tt>SN_2d</tt> is the SN per hour of 2D (15N, 1H) HSQC. =A= is constant, which has value of 0.8639 for (4,3) GFT HNNCABCA, 1.6019 for (4,3)D GFT CABCAcoNHN and 1.0153 for (4,3)D HABCABCONHN.

[[Image:Sn3.jpg]]

One can use [[NESG:UBNMR|UBNMR]] to run the measurement time predition by the following command: <tt>predict T2d SN2d Tc</tt> where

*<tt>T2d</tt> is the acquisition time of 2D [15N,1H] HSQC in hours
*<tt>SN2d</tt> is the SN distribution average of 2D [15N,1H] HSQC
*<tt>Tc</tt> is the rotational correlation time of protein in nanoseconds.

 

-- AlexEletski - 03 Mar 2008

← Older revision		Revision as of 15:47, 15 December 2009
Line 1:		Line 1:
	The minimum measurement time of (4,3)D GFT experiments can be reliably predicted from the S/N distribution of a 2D [15N, 1H] HSQC and the rotational correlation time tau_c. First, you need to generate an integrated peaklist.		The minimum measurement time of (4,3)D GFT experiments can be reliably predicted from the S/N distribution of a 2D [15N, 1H] HSQC and the rotational correlation time tau_c. First, you need to generate an integrated peaklist.

	==== '''Peak Integration in 2D [15N, 1H] HSQC''' ====		==== '''Peak Integration in 2D [15N, 1H] HSQC''' ====

	Having processed 2D (15N, 1H) HSQC as described in data process, one can do the peak picking and integration of 2D (15N, 1H) HSQC ~~manualy~~ or semi-~~automaticlly~~ as following:		Having processed 2D (15N, 1H) HSQC as described in data process, one can do the peak picking and integration of 2D (15N, 1H) HSQC manually or semi-automatically as following:

	*Peak picking and integration by using program XEASY		*Peak picking and integration by using program XEASY
Line 11:		Line 11:
	##Use <tt>in</tt> to automatic pick peaks with total peak number slightly more than expected by select adjust contour level; then manually remove side-chain amide peaks		##Use <tt>in</tt> to automatic pick peaks with total peak number slightly more than expected by select adjust contour level; then manually remove side-chain amide peaks
	##Use <tt>mn</tt> to measure noise level, and use <tt>tw</tt> to display the noise level value. Normally the value of 2.5 times of standard deviation ( ~250 if the noise level has been normalized ) is taken as noise level.		##Use <tt>mn</tt> to measure noise level, and use <tt>tw</tt> to display the noise level value. Normally the value of 2.5 times of standard deviation ( ~250 if the noise level has been normalized ) is taken as noise level.
	##Use <tt>ip</tt> to choose peak height (m) as integration mode, and use "ii" to integrate the whole spectrum. An [[NESG:XEASY\|XEASY]] external program "PeakintI" can be used to obtain more accurate peak height values, which is ~~discribed~~ below.		##Use <tt>ip</tt> to choose peak height (m) as integration mode, and use "ii" to integrate the whole spectrum. An [[NESG:XEASY\|XEASY]] external program "PeakintI" can be used to obtain more accurate peak height values, which is described below.
	##Use <tt>wp</tt> to save the peak list.		##Use <tt>wp</tt> to save the peak list.

Line 23:		Line 23:
	The SN distribution of resonances in a NMR spectra can be fit to the Gaussian distribution:		The SN distribution of resonances in a NMR spectra can be fit to the Gaussian distribution:

	[[Image:~~Sn_nhsqc~~.jpg]]		[[Image:Sn nhsqc.jpg]]

	<br><tt>f= aexp(-0.5((ln(SN_i)-ln(SN_0))/b)^2) </tt><br> where <tt>SN_0</tt> is the most populated S/N observed, <tt>f is the expected population at a ~~certern~~ SN value SN_i</tt>, <tt>a and b</tt> are ~~contants~~.		<br><tt>f= aexp(-0.5((ln(SN_i)-ln(SN_0))/b)^2) </tt><br> where <tt>SN_0</tt> is the most populated S/N observed, <tt>f is the expected population at a certain SN value SN_i</tt>, <tt>a and b</tt> are constants.

	The SN of NHSQC <tt>SN_0</tt> can be obtained as following:		The SN of NHSQC <tt>SN_0</tt> can be obtained as following:
Line 37:		Line 37:
	The SN distribution of resonances in other NMR spectra can also be fit to the Gaussian distribution as in 2D (15N, 1H) NHSQC:<br><br> <tt>f= aexp(-0.5((ln(SN_i)-ln(SN_0))/b)^2)</tt> <br><br> where <tt>SN_0</tt> is the most populated SN observed, <tt>f</tt> is the expected population at a certain SN value <tt>SN_i</tt>, <tt>a</tt> and <tt>b</tt> are constants. Based on this equation, one can calculate the expected SN_0 for a required peak detection yield.		The SN distribution of resonances in other NMR spectra can also be fit to the Gaussian distribution as in 2D (15N, 1H) NHSQC:<br><br> <tt>f= aexp(-0.5((ln(SN_i)-ln(SN_0))/b)^2)</tt> <br><br> where <tt>SN_0</tt> is the most populated SN observed, <tt>f</tt> is the expected population at a certain SN value <tt>SN_i</tt>, <tt>a</tt> and <tt>b</tt> are constants. Based on this equation, one can calculate the expected SN_0 for a required peak detection yield.

	Assuming that a peak shall have at least SN value of 2 in order to be observed or detected, and the average b for (4,3) GFT experiments is 0.8; if 95% peak detection yield is required, the ~~exptected~~ SN_0 is:<br><br><tt>SN_0=exp(ln2+1.644*b)=7.4 </tt><br><br>		Assuming that a peak shall have at least SN value of 2 in order to be observed or detected, and the average b for (4,3) GFT experiments is 0.8; if 95% peak detection yield is required, the expected SN_0 is:<br><br><tt>SN_0=exp(ln2+1.644*b)=7.4 </tt><br><br>

	[[Image:Sn2.jpg]]		[[Image:Sn2.jpg]]
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	[[Image:Sn3.jpg]]		[[Image:Sn3.jpg]]

	One can use [[NESG:UBNMR\|UBNMR]] to run the measurement time ~~predition~~ by the following command: <br><br><br> <tt>predict T2d SN2d Tc</tt><br>where		One can use [[NESG:UBNMR\|UBNMR]] to run the measurement time prediction by the following command: <br><br><br> <tt>predict T2d SN2d Tc</tt><br>where

	*<tt>T2d</tt> is the acquisition time of 2D [15N,1H] HSQC in hours		*<tt>T2d</tt> is the acquisition time of 2D [15N,1H] HSQC in hours

Estimation of measurement time - Revision history

Aringh at 15:47, 15 December 2009

Alex: Created page with 'The minimum measurement time of (4,3)D GFT experiments can be reliably predicted from the S/N distribution of a 2D [15N, 1H] HSQC and the rotational correlation time tau_c. First…'