Sequential HNCA

Ref.: A.Meissner, and O.W.Sørensen J.Magn.Reson.150,100-104 (2001). (A Sequential HNCA NMR Pulse Sequence for Protein Backbone Assignment)
Sequential HNCA pulse sequence
Delays: ' = S =  (2JNH)-1I = (2JNH)-1I; = gradient delay; TC = 7.6 ms, T = 22 ms, T' = 21 ms.
Phases for Varian Unity Inova:   = -y; {I = y; S = y - S } echo (t2 -> t3), {I = -y; S = -y - S } antiecho (t2 -> t3);
Phases for Bruker DRX:   = y; " = ; {I = -y; S = -y - S } echo (t2 -> t3), {I = y; S = y - S } antiecho (t2-> t3);
The uncompensated TROSY experiment employs SI = 0.
Gradient ratios: (G1 : G2 : G3) = (-7 : 3 : 1.987) echo; (G1 : G2 : G3) = (-8 : 2 : 3.013) antiecho.



Superposition of first t2 interferogram (echo) of sequential HNCA (interresidual correlations) and edited intra residual HNCA obtained by linear combination of HNCA TROSY and sequential HNCA in a ratio of 0.65 : -1. Sample: 4.18 mM 15N,13C-labeled CI2 21-83 (90% H2O/10% D2O, 25 deg. C, pH 4.2)

 


Varian:

exp s: sequential HNCA (default)
c: HNCA TROSY (first shaped CO  pulse is omitted and the second CO  pulse is simultaneously with the 1H and 15 pulse within the C evolution period. 

To generate a selective water pulse shape, adjust ref_pwr and ref_pw90 in the file $HOME/vnmrsys/shapelib/water_sc.inp according to your proton pulse calibration and type the command: Pbox -f water_sc.inp

An output like
>>> Set pulse width to 1.0000 ms <<<
>>> Set pulse power to 21 dB <<<
appears.

Remember to change the parameters pwater (pulse width in us) and pwaterlvl (pulse power).

For generating a selective CO  pulse adjust ref_pwr and ref_pw90 in the file $HOME/vnmrsys/shapelib/co_ib_off.inp according to your carbon pulse calibration. Adjust also the offset (ofs) to 120*dfrq. Run the command: Pbox -f co_ib_off.inp. change the parameters pwxcoilvl and pwxcoi according to the output. Apply the same procedure to the wurst2_co.inp file.
 

Processing of 3D sequential HNCA:

Data is States-TPPI in F1 and echo antiecho in F2. use your usual software for processing.

Installation:

For installation the compressed archive hnca_seq.tar.Z containing all components is required. Copy the file into your $HOME directory uncomress (uncompress hnca_seq.tar.Z) the file and unpack the archive (tar -xvf hnca_seq.tar). The files will be copied to the appropriate location within the vnmrsys directory. Load the parameters within vnmr ($HOME/vnmrsys/parlib/hnca_seq.par) and type the command seqgen(seqfil). Adjust Acquisition parameters (pulse lengths, offsets sweep widths, etc)

Program tested on: Varian Unity Inova, Vnmr6.1B, Pbox6.1C, Solaris7

Content
Pulse sequence:        hnca_seq.c
Parameters:            hnca_seq.par
Pbox input file:       water_sc.inp
                       ib_co_off.inp
                       wurst2_co.inp

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Bruker:

For installation the compressed archive hnca_seq.tar.gz containing all components is required. Copy the file to your spectrometer computer, uncomress (gunzip hnca_seq.tar.gz) the file and unpack the archive (tar -xvf hnca_seq.tar). Place gradient files trEA_1 (Gradient G1), trEA_2 (Gradient G2), and trEA_3 (Gradient G3) in the /u/exp/stan/nmr/lists/gp directory. (To change the ratio of the shaded gradients, these files can be edited. The two entries represent the relative strength for echo and antiecho, respectively). The gradient strengths are set within ased. All other files have to be located in the /u/exp/stan/nmr/lists/pp directroy.

Typical Parameters for DRX600 2D Version sequential H(N)CA

Processing of 3D seq_HNCA:

  • When encountering problems with the result of 3D FT please perform processing under consideration of Bug #2435 from Bruker Bug Database. (Apply the 'c' option with the tf3 command and choose equal 'xdim' values in F2 and F1)
Program tested on: Bruker DRX600, Xwinnmr2.6 patchlevel 4, Irix6.3

Content
Pulse sequence:           hnca_seq
                          hnca_seq2d
Gradient files:           trEA_1
                          trEA_2
                          trEA_3

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Document: Sequential HNCA (index.shtml)
Last modified: 2005-12-19