The 3D HMQC-TOCSY experiment is specifically designed
to obtain X-edited TOCSY spectra from which 1H-1H
TOCSY correlations can be clearly assigned even in overcrowded regions
(the mechanism involves an heteronuclear transfer via 1J(XH)
followed by a TOCSY step). Similar results can be obtained from an analog
3D HSQC-TOCSY 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 HMQC-TOCSY
experiment is usually applied on 13C-labeled proteins in D2O
and at natural abundance on concentrated samples. On the other hand, the
1H-15N HMQC-TOCSY is usually applied on 15N-labeled
molecules in H2O.
VERSIONS
The original 3D HMQC-TOCSY pulse sequence
(
89JMR634-84
and
92JMR433-99
)
basically consists of a 2D 1H-X HMQC pulse sequence followed
by an homonuclear 2D TOCSY pulse sequence.
Related experiments are:
The starting HMQC block could be replaced with other heteronuclear schemes
as, for instance, the HSQC pulse train (see, for instance, the 3D
HSQC-TOCSY experiment).
The TOCSY block can be replaced by a COSY
(
91JMR658-92
),
ROESY or a NOESY block (see, for instance, the 3D
HMQC-NOESY experiment).
The two building blocks can be reversed (see 3D
TOCSY-HMQC experiment)
A 2D version can be designed by fixing the variable 13C evolution
period in the HMQC block (2D w1-X-filtered TOCSY experiment) or
by fixing the variable 1H evolution period (see 2D
HMQC-TOCSY experiment). In addition, a 13C-selective 1D
HMQC-TOCSY could be obtained by fixing both variable evolution delays.
Several improved versions have been proposed incorporating the following
modifications:
Use of PFGs
(
91JMR664-94
).
In the absence of X-decoupling, the 3D HMQC-TOCSY experiment exhibits E.COSY
type cross-peaks, allowing the measurement of direct and long-range heteronuclear
coupling constants
(
91ANG1329
,
91JMR434-92
,
91JB403
, and
94JACS5823
).
A 3D DEPT-HMQC-TOCSY experiment has also been proposed
(
90ANG546
).
EXPERIMENTAL DETAILS
The 3D HMQC-TOCSY experiment can be recorded in automation
mode. Both blocks are usually optimized as in their 2D counterparts.
More details on practical implementation of the 3D HMQC-TOCSY experiment
on AVANCE spectrometers can be found in the corresponding Tutorial 3D
HMQC-TOCSY experiment
SPECTRA
The HMQC-TOCSY experiment affords a 3D spectrum in which X,
1H and 1H chemical shifts are displayed in three
independent dimensions. Thus, the F2(1H)-F3(1H)
plane of a 1H-15N HMQC-TOCSY spectrum will show a
typical 2D F1(NH)-F2(1H) TOCSY map. On
the other hand, the F2(1H)-F3(1H)
plane of a 1H-13C HMQC-TOCSY spectrum will show a
typical 2D F1(1H aliphatic)-F2(1H)
TOCSY map while the F1(13C)-F2(1H)
plane will correspond to a conventional 2D 1H-13C
HMQC spectrum.