Dear Tong Zhou,

NMR:

Nuclear magnetic resonance spectroscopy of proteins (usually abbreviated protein NMR) is a field of structural biology in which NMR spectroscopy is used to obtain information about the structure and dynamics of proteins, and also nucleic acids, and their complexes.

Advantages of NMR:

1. We can get several types of information from different types of experiments.

2. By NMR we can obtain angles, distances, coupling constants, chemical shifts, rate constants etc. These are really molecular parameters which could be examined more with computers and molecular modeling procedures.

3. With a suitable computer apparatus we can calculate the whole 3D structure

4. There are lots of possibilities to collect different data-sets from different types of experiments for the ability to resolve the uncertainties of one type of measurements

5. The motion of the segments (domains) can be examined.

6. NMR is capable to lead us for the observation of the chemical kinetics

7. Thermodynamic & kinetic data could be determined from a well-prepared (dynamic-) NMR experiment.

8. There is a possibility to investigate the influence of the dielectric constant, the polarity and any other properties of the solvent or some added material

 Disadvantages of NMR:

1. we have lots of atoms and a lot of extracted data from a system.

2. This is good for the more accurate determination of the structure, but not for the availability of higher molecular masses

3. The resolving power of NMR is less than some other type of experiments (e.g.: X-ray crystallography) since the information got from the same material is much more complex

4. The highest molecular mass which was examined successfully is just a 64kDa protein-complex

5. There are lots of cases when from a given data-set - a given type of experiment - we could predict two or more possible conformations, too

6. Unfortunately we are just able to determine the degree of probability of being of the protein segment in the given conformation

7. The cost of the experimental implementation is increasing with the higher strength and the complexity of the determination

X-ray:

To obtain X-ray data from a crystal, it needs to be placed in a monochromatic (single wavelength) X-ray beam. Subsequently, it is repeatedly exposed to the X-ray beam, while changing its orientation (usually rotating). Each exposure provides an image, similar to that shown above. Each spot on the image is a diffracted X-ray beam, which emerged from the crystal and was registered by the X-ray detector. Thousands of diffraction spots need to be collected to solve a protein structure. Depending on the type of the crystal (cell dimensions and symmetry), different strategies for data collection are followed and a different amount of data is collected. Usually the crystal is rotated in the X-ray beam one degree a time, and exposed to X-rays for a short period (seconds to minutes, depending on the intensity of the X-ray source). The intensities of these spots are subsequently used to calculate the electron density of the molecules within the crystal. The electron density, in turn, will tell us where the atoms are located, information which can be used to build a model of the molecule or molecules in the crystal.

Advantages of X-ray:

1. . We can examine also by this way the solvent effect since from different solvents the same protein may crystallize into different crystalloid form.

2. So we are able to force the protein to an other form of crystallization by the change of its solvent.

3.We could get the whole 3D structure by the systematic analysis of a good crystallized material

Disadvantages of X-ray:

1. The crystal structure is necessary only that proteins which can be crystallized are examinable

2. We cannot examine solutions and the behavior of the molecules in solution

3. This happens when we try to examine powders, gases

4. Studying of motions are not available

5. We can get only one parameter-set so we are able to observe only one conformation

6. There is no possibility to examine small parts in the molecule

7. There is no chance for direct determination of secondary structures and especially domain movements (big disadvantage against the NMR)

8. The hydrogen in the molecules are not examinable since it has only one electron

As a conclusion we must say that the two methods are very basic and critical in the field of protein structure determination. The strengths and weaknesses of one of the two methods fortunately are of those kinds which supplement the holes and gaps in the other method to make it possible that different kind of important data for a structural question can be answered with the parallel or supplemental application of the two methods. There may be several instances when only one method could be used. If the two methodologies had the same basics, principles, strengths and weaknesses we may be not capable to solve lots of structural problems. We should thank this possibility to the technical development which could catch different biophysical principles..

References

1. NMR spectroscopy

by Kurt D. Berndt

(http://iona.cryst.bbk.ac.uk/course/section8/ss-960531_22.html)       

  2.  NMR spectroscopy. Principles and applications

          by Henry Rzepa

          (http://www.ch.ic.ac.uk/local/organic/nmr.html)         

3. Structure determination of proteins with NMR spectroscopy

            by Horst Joachim Schirra

           (http://www.cryst.bbk.ac.uk/pps2/projects/schirra/html/home.htm)

          4. Magnetic Resonance

        by László Szilágyi (in hungarian), 1987,

          University of Lajos Kossuth (KLTE), Hungary

          5. The investigation of  ligand exchange processes of cyanide ion

            in the HCN-DCN and the Tl(edta)CN2- - CN-  systems by

            dynamic NMR spectroscopy (in hungarian)

            by Attila Ambrus,

            Diploma Thesis, 1996,  supervisor: István Bányai,

            Dept. of Physical Chemistry, University of Lajos Kossuth, Hungary

          6. X-ray Diffraction

           http://www-wilson.ucsd.edu/education/xraydiff/xraydiff.html)

          7. Diffraction methods (in hungarian)

           by Ferenc Joó,

           Dept. of Physical Chemistry, University of Lajos Kossuth, Hungary

          8. And some valuable conversations with an expert molecular modellist:

              Péter Bagossi

   http://www.cryst.bbk.ac.uk/pps97/assignments/projects/ambrus/html.htm

    Hoping this will be helpful,

Rafik

Hi there,

The main difference is that NMR requires a sample in solution whereas x-ray cristallography requires crystal sample. Therefore NMR determined structure gives the pic of the protein structure in solution, x-ray gives the picture of the protein structure within a crystal.

Dear Tong Zhou,

NMR:

Nuclear magnetic resonance spectroscopy of proteins (usually abbreviated protein NMR) is a field of structural biology in which NMR spectroscopy is used to obtain information about the structure and dynamics of proteins, and also nucleic acids, and their complexes.

Advantages of NMR:

1. We can get several types of information from different types of experiments.

2. By NMR we can obtain angles, distances, coupling constants, chemical shifts, rate constants etc. These are really molecular parameters which could be examined more with computers and molecular modeling procedures.

3. With a suitable computer apparatus we can calculate the whole 3D structure

4. There are lots of possibilities to collect different data-sets from different types of experiments for the ability to resolve the uncertainties of one type of measurements

5. The motion of the segments (domains) can be examined.

6. NMR is capable to lead us for the observation of the chemical kinetics

7. Thermodynamic & kinetic data could be determined from a well-prepared (dynamic-) NMR experiment.

8. There is a possibility to investigate the influence of the dielectric constant, the polarity and any other properties of the solvent or some added material

 Disadvantages of NMR:

1. we have lots of atoms and a lot of extracted data from a system.

2. This is good for the more accurate determination of the structure, but not for the availability of higher molecular masses

3. The resolving power of NMR is less than some other type of experiments (e.g.: X-ray crystallography) since the information got from the same material is much more complex

4. The highest molecular mass which was examined successfully is just a 64kDa protein-complex

5. There are lots of cases when from a given data-set - a given type of experiment - we could predict two or more possible conformations, too

6. Unfortunately we are just able to determine the degree of probability of being of the protein segment in the given conformation

7. The cost of the experimental implementation is increasing with the higher strength and the complexity of the determination

X-ray:

To obtain X-ray data from a crystal, it needs to be placed in a monochromatic (single wavelength) X-ray beam. Subsequently, it is repeatedly exposed to the X-ray beam, while changing its orientation (usually rotating). Each exposure provides an image, similar to that shown above. Each spot on the image is a diffracted X-ray beam, which emerged from the crystal and was registered by the X-ray detector. Thousands of diffraction spots need to be collected to solve a protein structure. Depending on the type of the crystal (cell dimensions and symmetry), different strategies for data collection are followed and a different amount of data is collected. Usually the crystal is rotated in the X-ray beam one degree a time, and exposed to X-rays for a short period (seconds to minutes, depending on the intensity of the X-ray source). The intensities of these spots are subsequently used to calculate the electron density of the molecules within the crystal. The electron density, in turn, will tell us where the atoms are located, information which can be used to build a model of the molecule or molecules in the crystal.

Advantages of X-ray:

1. . We can examine also by this way the solvent effect since from different solvents the same protein may crystallize into different crystalloid form.

2. So we are able to force the protein to an other form of crystallization by the change of its solvent.

3.We could get the whole 3D structure by the systematic analysis of a good crystallized material

Disadvantages of X-ray:

1. The crystal structure is necessary only that proteins which can be crystallized are examinable

2. We cannot examine solutions and the behavior of the molecules in solution

3. This happens when we try to examine powders, gases

4. Studying of motions are not available

5. We can get only one parameter-set so we are able to observe only one conformation

6. There is no possibility to examine small parts in the molecule

7. There is no chance for direct determination of secondary structures and especially domain movements (big disadvantage against the NMR)

8. The hydrogen in the molecules are not examinable since it has only one electron

As a conclusion we must say that the two methods are very basic and critical in the field of protein structure determination. The strengths and weaknesses of one of the two methods fortunately are of those kinds which supplement the holes and gaps in the other method to make it possible that different kind of important data for a structural question can be answered with the parallel or supplemental application of the two methods. There may be several instances when only one method could be used. If the two methodologies had the same basics, principles, strengths and weaknesses we may be not capable to solve lots of structural problems. We should thank this possibility to the technical development which could catch different biophysical principles..

References

1. NMR spectroscopy

by Kurt D. Berndt

(http://iona.cryst.bbk.ac.uk/course/section8/ss-960531_22.html)       

  2.  NMR spectroscopy. Principles and applications

          by Henry Rzepa

          (http://www.ch.ic.ac.uk/local/organic/nmr.html)         

3. Structure determination of proteins with NMR spectroscopy

            by Horst Joachim Schirra

           (http://www.cryst.bbk.ac.uk/pps2/projects/schirra/html/home.htm)

          4. Magnetic Resonance

        by László Szilágyi (in hungarian), 1987,

          University of Lajos Kossuth (KLTE), Hungary

          5. The investigation of  ligand exchange processes of cyanide ion

            in the HCN-DCN and the Tl(edta)CN2- - CN-  systems by

            dynamic NMR spectroscopy (in hungarian)

            by Attila Ambrus,

            Diploma Thesis, 1996,  supervisor: István Bányai,

            Dept. of Physical Chemistry, University of Lajos Kossuth, Hungary

          6. X-ray Diffraction

           http://www-wilson.ucsd.edu/education/xraydiff/xraydiff.html)

          7. Diffraction methods (in hungarian)

           by Ferenc Joó,

           Dept. of Physical Chemistry, University of Lajos Kossuth, Hungary

          8. And some valuable conversations with an expert molecular modellist:

              Péter Bagossi

   http://www.cryst.bbk.ac.uk/pps97/assignments/projects/ambrus/html.htm

    Hoping this will be helpful,

Rafik