CAN for Bruker - pulse sequence to correlate CA and N (Wagner Lab)

From NMR Wiki

Basic facts about n_can_2h_ct_2d_ul.kt

Synopsis: CAN - 15N detected pulse sequence to correlate amide Ni with Cαi and Cαi-1

Family: CAN

Sample requirements: Uniformly D,13C,15N labeled

Code authors:  Gerhard Wagner,  Koh Takeuchi

Copyright: Harvard Medical School

Hardware requirements:  HCND probe, optimized for 15N and 13C detection

Introduction

Please see CAN concept document for more explanation of this experiment.

Delays

• d1 - recycling delay [1.6s]
• d16 - gradient recovery delay [200μs]

Constants

• cnst21 - average chemical shift of CO atoms [176]
• cnst22 - average chemical shift of CA atoms in ppm [55]

Acquisition dimensions

• aq - acquisition time in directly detected ¹⁵N dimension

Carrier frequencies

• o1p - ¹⁵N - center of amide nitrogen region
• o2p - ¹H - water resonance ~4.7 ppm
• o3p - ¹³C - center of Cα region [55ppm]
• o4p - ²H - water resonance ~4.7 ppm

Hard pulses

• p1 - length of the hard 90^o pulse for ¹⁵N
• pl1 - power level for p1 pulse

Gradients

• p16 - length of the gradient pulse [1ms]
• gpnam1 - name of the gradient waveform [SINE.100]
• gpz1 - strength of z gradient - 10 G/cm ~[20]%
• gpx1 - 0
• gpy1 - 0
• d16 - gradient recovery delay [200μs]

Shaped pulses

Power level for the shaped pulses needs to be calibrated with the stdisp tool.

There are five shaped pulses in this pulse sequence: sp15, sp23, sp24, sp25 and sp26.

Offsets for the shaped pulses are 0 Hz except sp26 and sp15

sp15

This pulse is used as a part of the cpd3 (mlevsp180) decoupling program

The waveform employed is wurst frequency swept adiabatic inversion pulse.

Frequency sweep - 40 kHz, duration 2ms - specific to 500 MHz instrument.

The purpose of this pulse is to simultaneously decouple CO and CA.

• pcpd3 - length 2ms
• pl16 - power level - not sp15 - because this pulse is a part of composite pulse decoupling program
• spnam15 - waveform wurst-40k-2ms.kt - included with this record
• spoffs15 - offset for the pulse sp15 - 115.5 ppm - the middle of CO and CA chemical shift

sp23

90^o excitation pulse for ¹³C.

• p11 - length [320μs]
• sp23 - power level
• spnam23 - waveform Q5.1000

sp24 and sp26

180^o inversion pulse for ¹³C.

• p12 - length [256μs]
• sp24, sp26 - power levels
• spnam24, spnam26 - waveform Q3.1000
• spoffs26 - offset for pulse sp26 - center of carbonyl (CO) region in Hertz

sp25

180o refocussing pulse for 13C.

• p11 - length [320μs]
• sp25 - power level
• spnam25 - waveform - Q5tr.1000

Decoupling

Default lengths of decoupling given here are for 500 MHz instrument.

cpd2

Used to decouple ¹H - protons.

• pl12 - power level
• pcpd2 - duration of the 90^o pulse for waltz16 [80μs]
• cpdprg2 - decoupling program waltz16

cpd3

See setup for the shaped pulse sp15 as mlevsp180 program uses shaped pulse sp15

• cpdprg3 - program for decoupling [mlevsp180]
• pl16 - power level used by the waveform assigned to spnam15

cpd4

Deuterium (²H) decoupling.

• pl17 - power level for the 90^o within the decoupling program
• pcpd4 - length of the 90^o within the decoupling program [250μs]
• cpdprg4 - program for decoupling [waltz16]

References

1. Takeuchi, K and Heffron, G and Sun, ZYJ and Frueh, DP and Wagner, G. Nitrogen-detected CAN and CON experiments as alternative experiments for main chain NMR resonance assignments. Journal of Biomolecular NMR pp. 1--12. BibTeX [cancon]