Nuclear Magnetic Resonance (NMR) spectroscopy represents a fundamental source of information and insight into molecular structure and intermolecular interactions. The high field 700 MHz spectrometer located in the Life Science Building at York University provides excellent sensitivity and resolution and is routinely applied for the detection and study of a wide variety of large and small molecules in solution. The structure and behaviour of proteins, nucleic acids, and other polymers can be routinely investigated as well as medium size and small molecule systems. This state-of-the-art spectrometer benefits from the inclusion of cryoprobe technology and fast acquisition times using NonUniform Sampling (NUS) software methods.
The Life Science NMR Facility at York is engaged with understanding the causes of many physiological disorders. The comprehensive and diverse nature of these studies is explored through NMR studies of protein, nucleic acid and other intracellular systems. Protein structure and binding interactions detected by NMR provides important and unique information on a molecular scale. Even larger proteins can be studied by the introduction of deuterium labelling. Protein behaviour and substrate interactions are typically observed through the determination of interaction properties. Kinetic information of enzyme-substrate interactions can routinely followed by 2D 1H-15N HSQC while enzymatic reactions can accurately be measured by 1D 1H experiments. Ongoing NMR studies are continually revealing new insight into biomolecular interactions.
The 700 MHz NMR spectrometer is equipped with the following features:
- Optimized for biological and small molecule data acquisition
- 5 mm (or smaller) sample tubes
- Can observe or decouple 1H, 13C, 15N, 2H
- Sensitivity enhanced Cryoprobe equipped with x,y,z pulsed field gradients
- Variable temperature operation
- TOPSPIN 3.1 software
- Topshim software for rapid optimization of magnetic field homogeneity
- Multidimensional 1D, 2D, 3D, … nD data acquisition.
Protein structure determination is facilitated through the liquid helium cryoprobe capable of observation of 1H, 13C,15N, and 2H decoupling for 3D to nD pulsed field gradient experiments. The addition of non-uniform sampling acquisition and processing software greatly reduces data collection time.
The NMR Facility at York University offers a wide range of NMR services from the routine acquisition of 1D proton spectra to multidimensional data for protein structure determination. Customers may choose to submit samples for analysis or may arrange for NMR training to enable frequent data acquisition at convenient times. Consultation services are also available to ensure that the most informative data can be collected. Detailed analysis reports can be completed by request.
For sample submission please complete the following form:
NMR Request Form-700
Biological NMR Data:
- High Resolution (Solution) NMR
- Data Acquisition 1D, 2D, 3D…
- Other nuclei
Small Molecule Analysis
High Resolution (Solution) NMR
- Data Acquisition
- microgram quantities
- 1D, 2D, 3D…
- 1H, 13C, 15N assignments
- Structure Elucidation
- Quantitative NMR
- Other nuclei
NMR schedules will cover 2 week periods. All NMR time requests must be submitted 1 week in advance of the start date. For example: for the period from Monday March 3 to Sunday March 16, 2014, requests must be received by Monday Feb 24. Preferred days and times should be included in the submission. Upon evaluation of the requests, the researchers will be notified of their allotted time so that he/she may reserve it online using FACES.
Time allocated will be determined by the total time requested by all researchers. In the event of an abnormally large time request, time will be allocated evenly between the researchers submitting the request. For example, if 3 people request large blocks of time, the maximum allotted time will be 33% per researcher for the 2 week period or 4.7 days.
In the event that long blocks of time are required by a single researcher, a reasonable effort will be made to arrange for time to be continuous (eg. finish one 2 week block and begin the next 2 week block).
In the event that the sample does not give acceptable spectra (eg. decomposition or unexpected impurities) the researcher can release the reserved the time and notify Howard.
Switching scheduled time can only be done by mutual agreement between researchers.
Lost time due to instrument repair will be rescheduled to a later time so that only the researcher with time scheduled for that day will be affected.
Nitrogen fills will be completed Thursday afternoon or Friday during the day. Helium fills will also be completed every few months.
In the early years of 2D NMR, the organic chemist’s dream of the perfect 2D NMR experiment for structure elucidation was a 2D 13C-13C correlation. From this idea evolved the Incredible Natural Abundance Double Quantum Transfer Experiment (INADEQUATE). Until recently, the INADEQUATE experiment was also the insensitive experiment due to the low natural abundance and long T1 relaxation times for 13C nuclei. The natural abundance of 13C is 1.1% and the chance of having two 13C nuclei directly bonded together is 1.1% of 1.1% or 0.0121% out of all adjacent carbon atoms in a molecule. Therefore from a pure chemical dissolved in the NMR solvent, only 0.0121% would give a 13C-13C correlation. The other main problem of the 2D 13C-13C INADEQUATE experiment is the long T1 relaxation time of the 13C nuclei. Typically 13C nuclei with attached protons relax faster than nonprotonated 13C nuclei. As a result, the 1D 13C NMR spectra usually show the lowest intensity for quaternary carbons. Ideally, to get the full intensity from slowly relaxing nuclei, a period of 5 x T1 is required for the nuclei to completely relax. In reality, this long delay is rarely used and the spectrum is collected with a shorter relaxation delay. The intensity of the correlations in the 2D INADEQUATE spectrum will be reduced if the relaxation delay between scans is too short and a lengthened delay can result in longer acquisition times. The 700 MHz ultra high field NMR features the addition of a cryogenically cooled detection coil in the Cryoprobe™, resulting in superior quality spectra obtained from far less sample than ever before.
The slide show shows the assignment process from identification of the methoxy- carbon-13 resonance to the use of the 2D INADEQUATE beginning with methyl carbon connectivity.