/
T1N15_tr.cw
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T1N15_tr.cw
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; 15N-T1 relaxation experiment with TROSY read-out
; for 15N, 15N13C, 2H15N and 2H15N13C labelled proteins
; written by NL 10/25/11
; see footnotes
#include <Avance.incl>
#include <Grad.incl>
#include <Delay.incl>
;#define LABEL_CN ; switch on for 13C labelled samples
; ALSO SET 13C DECOUPLING (AT TOP, 77, AND BOTTOM OF PROGRAM)
#define TEMP_COMPENSATION
"in0=inf1*0.5"
# ifdef LABEL_CN
"d0=97u-p4*2+p7*0.66-p1*0.5"
"d25=20m-p15*0.5-p4*4-8u"
#else
"d0=100u+p7*0.66-p1*0.5"
"d25=20m-p15*0.5"
#endif /*LABEL_CN*/
"d11=30m"
"DELTA=2.65m"
"DELTA1=2.65m"
"DELTA2=2.65m-p22-p11-300u"
"DELTA3=2.65m-p23-p10-300u"
"DELTA4=260u-p24-p1*0.66"
"d27=p24+35u"
"l1=1"
"l2=1"
"cnst21=176"
"cnst22=56"
"cnst18=-800"
"spoff4=bf2*((cnst22-cnst21)/1000000)"
1 ze
1m
2 d11 ;do:f2 ; 13C DECOUPLING SWITCH OFF!
1m LOCKH_OFF
3m
3 1m
1m
4 3m
5 2m BLKGRAD
10u pl1:f1
10u ;pl4:f2 ; 13C POWER
10u pl7:f3
(p7 ph0):f3
10u
;---------temperature compensation and d1 recovery delay---------
# ifdef TEMP_COMPENSATION
"d17=d1-p18"
10u fq=cnst18(bf ppm):f3
10u pl8:f3
(p18 ph0):f3 ; 15N pulse is applied far off-resonance
10u
10u fq=0:f3
d17
# else
d1
# endif
1m UNBLKGRAD
10u pl7:f3
;------- kill steady state 15N ------------
(p7 ph0):f3
5u
p20:gp6
200u
;------- first INEPT Hz-> 2HxNz -----------
(p1 ph0):f1
5u
DELTA gron0 ; soft gradient to prevent radiation damping
5u groff
(center(p1*2 ph0):f1 (p7*2 ph0):f3)
5u
DELTA gron0
5u groff
;------- rephase 2HxNz to Nz------ --------
(p1 ph5):f1 (p7 ph0):f3
5u
DELTA1 gron1 ; soft gradient to prevent radiation damping
5u groff
(center (p1*2 ph0):f1 (p7*2 ph0):f3)
5u
DELTA1 gron1
5u groff
(p7 ph6):f3 ; phase-cycle Nz, -Nz for Freeman-Hill decay
5u
;--------------------------------------------
(p1 ph2):f1 ; purge pulse to kill any residual HzNz
5u
p21:gp7 ; cleaning gradient
100u
100u pl0:f1
;------15N T1 relaxation period--------------
if "l2==1" goto 77 ; jump to 77 for first relaxation data point, needs to be 0 in vclist
70 d25*0.5
# ifdef LABEL_CN
3u pl4:f2
(p4*2 ph0 3u 3u pl2 p4*2:sp4 ph0):f2
# endif
d25*0.5
(p15:sp5 ph0):f1
d25*0.5
# ifdef LABEL_CN
3u pl4:f2
(p4*2 ph0 3u 3u pl2 p4*2:sp4 ph0):f2
# endif
d25*0.5
lo to 70 times c ; delay=c*2*d25 (20ms)
;------Echo/ Anti-echo encoding for TROSY read-out------------
77 3u
3u ;pl4:f2 ; 13C POWER
3u pl1:f1
if "l1==1"
{
(p7 ph7):f3
10u
p25:gp5
200u
(p7*2 ph7):f3
10u
p25:gp5*-1
}
else
{
(p7 ph17):f3
10u
p25:gp5*-1
200u
(p7*2 ph17):f3
10u
p25:gp5
}
;------ t1 (15N) evolution period ------------------------------
d0
# ifdef LABEL_CN
(p4*2 ph0 3u 3u pl2 p4*2:sp4 ph0):f2
# endif
d0
;------ start TROSY read-out------------------------------------
if "l1==1"
{
(p1 ph1):f1 ; Echo
3u
3u pl0:f1
(p11:sp11 ph11:r):f1
6u
}
else
{
(p1 ph3):f1 ; Anti-Echo
3u
3u pl0:f1
(p11:sp11 ph13:r):f1
6u
}
5u pl1:f1
;goto 999 ; optimization of water supression
DELTA2
p22:gp2
300u
(center (p1*2 ph0):f1 (p7*2 ph0):f3)
7u
p22:gp2
DELTA2
300u pl0:f1
;-------------------------------------------------
(p11:sp12 ph12:r):f1
5u
3u pl1:f1
if "l1==1"
{
(p1 ph0):f1 (p7 ph1):f3 ; Echo
}
else
{
(p1 ph0):f1 (p7 ph3):f3 ; Anti-Echo
}
;goto 999 ; for optimization of water supression
DELTA3
p23:gp3
200u
100u pl10:f1
(center(p10 ph10:r 5u pl1 p1*2 ph0 5u pl10 p10 ph10:r):f1 (p7*2 ph0 d27):f3)
5u
;goto 999 ; for optimization of water supression
p23:gp3
DELTA3
DELTA4
(p7 ph0):f3
5u
p24:gp4 ; Echo/Anti-echo decoding gradient
999 5u
5u ;pl31:f2 ; 13C DECOUPLING POWER
20u BLKGRAMP
go=2 ph31 ;cpds2:f2 ; 13C DECOUPLING
1m ;do:f2 ; 13C DECOUPLING SWITCH OFF!
1m LOCKH_OFF
d11 wr #0 if #0 zd
1m ivc
1m iu2
lo to 3 times l6
1m iu1
1m igrad EA
1m ru2
lo to 4 times 2
1m id0
1m ru1
lo to 5 times l3
1m
1m BLKGRAD
exit
ph0=0
ph1=1
ph2=2
ph3=3
ph5=1
ph6=1 1 1 1 3 3 3 3
ph10=2
ph11=3
ph12=0
ph13=1
ph7=1 0 3 2
ph17=1 2 3 0
ph31=1 2 3 0 3 0 1 2
;-------------NOTES----------------------
;o1p = 4.7 ppm
;o2p=176 ppm (CO)
;o3p=119 ppm
;NS=8*n
;in0=inf/2
;SW=1/(2*in0)
;echo-antiecho in N15 (process as Complex in NmrDraw before splitting the spectra)
; loop counters
;l3: number of complex points (td1 / 2)
;l6: number of relaxation points
; 1H pulses
;p1: 90 deg hard 1H pulse @pl1
;pl1: 1H 90 deg
;pl0: 120 dB
;p10: 1000u (@ 700 MHz) 90 deg soft rectangular water flip-back pulse (pl10)
;p11: 1600u (@ 700 MHz) 90 deg Sinc1.1000 water flip-back pulse (sp11,sp12)
;p15: 1700u (@ 700 MHz) 180 deg IBurp2 pulse on 1H (sp5)
;sp5: 180 deg IBurp2 pulse on 1H (p15)
;sp11: 90 deg Sinc1.1000 water flip-back pulse
;sp12: 90 deg Sinc1.1000 water flip-back pulse
;spnam5: IBurp2
;spnam11: Sinc1.1000
;spnam12: Sinc1.1000
;spoffs5: 2730Hz @ 700 MHz (8.6 ppm) , should be centered in amide region but not touch the water
; 13C pulses
;p4: 13CO selective 90 deg (20.31us @ 700 MHz) @pl4
;pl2: 120 dB
;pl4: 13C 90 deg
;sp4: 13CA selective 90 deg (p4)
;CPDPRG2: garp (aq C' decoupling)
;pcpd5: C' decoupling (140u or 280u @pl31)
;pl31: C' decoupling power
;15N pulses
;p7 : 90 deg hard 15N pulse @pl7
;p18 : maximum duration of spin-lock; temperature compensation
;pl7 :15N 90 deg
;pl8: 15N spin-lock power
; gradients
;p20: 1000u
;p21: 200u
;p22: 300u
;p23: 1000u
;p24: 60.8u Echo/Anti-echo decoding gradient
;p25: 300u Echo/Anti-echo half-encoding gradient
;for z-only gradients
;gpz0: 3%
;gpz1: 2%
;gpz2: 10%
;gpz3: 50%
;gpz4: 33%
;gpz5: -33%
;gpz6: 30%
;gpz7: -50%
;gpnam2 SINE.10
;gpnam3 SINE.50
;gpnam4 SINE.10
;gpnam5 SINE.10
;gpnam6 SINE.10
;gpnam7 SINE.10