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 =  (2J_NH)^-1; _I = (2J_NH)^-1 - _I; = gradient delay; T_C = 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 _S = _I = 0.
Gradient ratios: (G1 : G2 : G3) = (-7 : 3 : 1.987) echo; (G1 : G2 : G3) = (-8 : 2 : 3.013) antiecho.

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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)

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 15N  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)

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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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)

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

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