The 3D TOCSY-HSQC experiment is specifically designed
to obtain X-edited TOCSY spectra of X-labeled biomolecules from which homonuclear
TOCSY correlations can be clearly assigned even in overcrowded regions
(the mechanism involves a TOCSY step followed by an heteronuclear transfer
via 1J(XH)). Similar results can be obtained from an analog
3D TOCSY-HMQC experiment.
REQUIREMENTS
Implementation on AVANCE spectrometers is feasible. Improved
versions using pulsed field gradients (PFGs) are also available and, therefore,
gradient technology is required. The 1H-13C TOCSY-HSQC
experiment is usually applied on 13C-labeled proteins in D2O
and the 1H-15N TOCSY-HSQC is usually applied on 15N-labeled
molecules in H2O.
VERSIONS
The original TOCSY-HSQC pulse sequence basically consists of
an homonuclear 2D TOCSY pulse sequence followed by a 2D 1H-X
HSQC pulse sequence. However, the HSQC block could be replaced with
other schemes: decoupled HSQC, CT-HSQC, PEP-HSQC, HMQC ...(see, for instance,
the 3D TOCSY-HMQC experiment).
Some considerations:
Qualitative J(HA,HB) values can be extracted from the relative intensities
in a 1H-15N experiment.
3D HCCH-type experiments give better results than the 1H-13C
experiment.
Several improved versions have been proposed incorporating the following
modifications:
Improved sensitivity incorporating the PEP methodology combined with PFGs
(
96JB319
).
Simultaneous evolution of 13C and 15N chemical shift
in the t2 period
(
94BIO14858
).
Use of TROSY approach
The initial TOCSY block can be replaced by other homonuclear mixing processes
as described in the 3D TOCSY-NOESY-HSQC and NOESY-TOCSY-HSQC experiments
(
92JMR408-96
).
EXPERIMENTAL DETAILS
The 3D TOCSY-HSQC experiment can be recorded in automation
mode. More details on practical implementation of the 3D TOCSY-HSQC experiment
on AVANCE spectrometers can be found in the corresponding Tutorial 3D
TOCSY-HSQC experiment
SPECTRA
The TOCSY-HSQC experiment affords a 3D spectrum in which
1H, X and 1H chemical shifts are displayed in three
independent dimensions. Thus, the F1(1H)-F3(1H)
plane of a 1H-15N TOCSY-HSQC spectrum will show a
typical 2D F1(1H)-F2(NH) TOCSY map. On
the other hand, the F1(1H)-F3(1H)
plane of a 1H-13C TOCSY-HSQC spectrum will show a
typical 2D F1(1H)-F2(1H aliphatic)
TOCSY map while the F1(1H)-F2(13C)
plane will correspond to a conventional 2D 1H-13C
HSQC spectrum.
