With reference to the comment by Sheng:
One of bottle neck in protein solution structure study by
NMR is that we need quite concentrated protein in water.
Now this bottle neck has been enlarged to quite extent due
to the gradient technology. We might be still doing protein
'conditioning' on the protein that I currently study if we did
not have the gradient. Our receiver gain is normally set at
around 512 to 1k for the protein in water solution without
gradient. Now we can set receiver gain to 16K. Using
gradient together with facelift program I now can get pretty
good spectrum of low concentration protein otherwise
beyond my reach.
.... perhaps someone could help?
I'm currently attempting to accumulate worthwhile NOE
data on a particularly unhelpful small (6k) Gla
containing protein. Metal incorporation is required
for the establishment of tertiary structure, which does
not help. The limitation on protein concentration is some
3.4mg/ml, so you see the problem.
Regardless of presat strategies (3-9-19 watergate etc. etc.)
yep, RG with gradients is way up. What I have found (despite
many weeks of optimisation) is that the resolution with
gradients (z only) is terrible. S/N requires that gradients
are used however, so I have no way out.
Current pulprog (since I've been asked..)
;noesygstp19
;avance-version
;2D homonuclear correlation via dipolar coupling
;dipolar coupling may be due to noe or chemical exchange.
;phase sensitive using TPPI
;water suppression using 3-9-19 pulse sequence with gradients
;M. Piotto, V. Saudek & V. Sklenar, J. Biomol. NMR 2, 661 - 666 (1992)
;V. Sklenar, M. Piotto, R. Leppik $ V. Saudek, J. Magn. Reson.,
; Series A 102, 241 -245 (1993)
#include <Avance.incl>
#include <Grad.incl>
;;d0=3u
;;d12=20u
1 ze
2 d1
3 d12 pl1:f1
p1 ph1
d0
p1 ph2
d8
50u UNBLKGRAD
GRADIENT(cnst21)
d16
p1 ph3
d12 pl18:f1
GRADIENT(cnst22)
d16
p28*0.231 ph4
d19*2
p28*0.692 ph4
d19*2
p28*1.462 ph4
d19*2
p28*1.462 ph5
d19*2
p28*0.692 ph5
d19*2
p0*0.231 ph5
46u
GRADIENT(cnst23)
d16
4u BLKGRAD
go=2 ph31
d1 wr #0 if #0 id0 ip1 zd
lo to 3 times td1
exit
ph1=0 2
ph2=0 0 0 0 0 0 0 0 2 2 2 2 2 2 2 2
ph3=0 0 2 2 3 3 1 1
ph4=0
ph5=2
ph31=0 2 2 0 1 3 3 1 2 0 0 2 3 1 1 3
;pl1 : f1 channel - power level for pulse (default)
;pl18: f1 channel - power level for 3-9-19-pulse (watergate)
;p0 : f1 channel - 90 degree pulse at pl18
; use for fine adjustment
;p1 : f1 channel - 90 degree high power pulse
;p2 : f1 channel - 180 degree high power pulse
;p16: homospoil/gradient pulse
;p28: f1 channel - 90 degree pulse at pl18
;d0 : incremented delay (2D) [3 usec]
;d1 : relaxation delay; 1-5 * T1
;d8 : mixing time
;d12: delay for power switching [20 usec]
;d16: delay for homospoil/gradient recovery
;d19: delay for binomial water suppression
; d19 = (1/(2*d)), d = distance of next null (in Hz)
;in0: 1/(1 * SW) = 2 * DW
;nd0: 1
;NS: 8 * n
;DS: 16
;td1: number of experiments
;MC2: TPPI
;use gradient program (GRDPROG) : 2sine
;use gradient ratio: cnst21 : cnst22
; 1 : 1
Can someone comment on the problems.... Many thanks.
,,,
(o o)
+--------------------------oOO--(_)---OOo----------------------------+
| +----------------------------------------------------------------+ |
| | | |
| | Dr. Grahame Harden | |
| | | |
| | Department of Chemistry | |
| | | |
| | University of Oulu, Linnanmaa, SF-90570, Finland | |
| | | |
| | gharden@phoenix.oulu.fi | |
| +----------------------------------------------------------------+ |
+--------------------------------------------------------------------+