Heteronuclear single quantum correlation
Encyclopedia
The Heteronuclear Single Quantum Coherence (HSQC) experiment is used frequently in NMR
spectroscopy of organic molecules and is of particular significance in the field of protein NMR. It was invented by Geoffrey Bodenhausen and D. J. Ruben in 1980. The resulting spectrum is two-dimensional with one axis for 1H and the other for a heteronucleus (an atomic nucleus
other than a proton), most often 13C
or 15N. The spectrum contains a peak for each unique proton attached to the heteronucleus being considered. Thus, if the chemical shift
of a specific proton is known, the chemical shift of the coupled heteronucleus can be determined, and vice versa.
, or using isotopically labeled protein. The latter can be recorded on much lower concentrations of protein, but requires recombinant expression of the protein.
Each residue
of the protein
(except proline
) has an amide proton attached to a nitrogen
in the peptide bond
. If the protein is folded, the peaks are usually well dispersed, and most of the individual peaks can be distinguished. Being a relatively cheap and quick experiment, the HSQC is useful to screen candidates for structure
determination by NMR. The number of peaks in the spectrum should match the number of residues in the protein (though sidechains with nitrogen-bound protons will add additional peaks). It will probably be difficult to solve the structure of the protein if this is not the case. The labour-intensive process of structure determination is usually not undertaken until a good HSQC is obtained.
It is not possible to assign the HSQC spectrum by itself, or in other words to determine which peaks correspond to a particular residue in the protein. This process can be done in different ways as outlined in the protein NMR article. The assignment of the spectrum is usually the first step in a structure determination, and is essential for a meaningful interpretation of more advanced NMR experiments.
The HSQC experiment is also useful for detecting interactions with ligand
s, such as other proteins or drugs. By comparing the HSQC of the free protein with the one bound to the ligand, it is possible to find the changes in the chemical shift
s of the peaks, which is most likely to occur in the binding interface.
NMR
NMR may refer to:Applications of Nuclear Magnetic Resonance:* Nuclear magnetic resonance* NMR spectroscopy* Solid-state nuclear magnetic resonance* Protein nuclear magnetic resonance spectroscopy* Proton NMR* Carbon-13 NMR...
spectroscopy of organic molecules and is of particular significance in the field of protein NMR. It was invented by Geoffrey Bodenhausen and D. J. Ruben in 1980. The resulting spectrum is two-dimensional with one axis for 1H and the other for a heteronucleus (an atomic nucleus
Atomic nucleus
The nucleus is the very dense region consisting of protons and neutrons at the center of an atom. It was discovered in 1911, as a result of Ernest Rutherford's interpretation of the famous 1909 Rutherford experiment performed by Hans Geiger and Ernest Marsden, under the direction of Rutherford. The...
other than a proton), most often 13C
Carbon-13
Carbon-13 is a natural, stable isotope of carbon and one of the environmental isotopes. It makes up about 1.1% of all natural carbon on Earth.- Detection by mass spectrometry :...
or 15N. The spectrum contains a peak for each unique proton attached to the heteronucleus being considered. Thus, if the chemical shift
Chemical shift
In nuclear magnetic resonance spectroscopy, the chemical shift is the resonant frequency of a nucleus relative to a standard. Often the position and number of chemical shifts are diagnostic of the structure of a molecule...
of a specific proton is known, the chemical shift of the coupled heteronucleus can be determined, and vice versa.
HSQC in protein NMR
The 15N HSQC experiment is probably the most frequently recorded experiment in protein NMR. The HSQC experiment can be performed either using the natural abundance of the 15N isotopeIsotope
Isotopes are variants of atoms of a particular chemical element, which have differing numbers of neutrons. Atoms of a particular element by definition must contain the same number of protons but may have a distinct number of neutrons which differs from atom to atom, without changing the designation...
, or using isotopically labeled protein. The latter can be recorded on much lower concentrations of protein, but requires recombinant expression of the protein.
Each residue
Residue (chemistry)
In chemistry, residue is the material remaining after a distillation or an evaporation, or to a portion of a larger molecule, such as a methyl group. It may also refer to the undesired byproducts of a reaction....
of the protein
Protein
Proteins are biochemical compounds consisting of one or more polypeptides typically folded into a globular or fibrous form, facilitating a biological function. A polypeptide is a single linear polymer chain of amino acids bonded together by peptide bonds between the carboxyl and amino groups of...
(except proline
Proline
Proline is an α-amino acid, one of the twenty DNA-encoded amino acids. Its codons are CCU, CCC, CCA, and CCG. It is not an essential amino acid, which means that the human body can synthesize it. It is unique among the 20 protein-forming amino acids in that the α-amino group is secondary...
) has an amide proton attached to a nitrogen
Nitrogen
Nitrogen is a chemical element that has the symbol N, atomic number of 7 and atomic mass 14.00674 u. Elemental nitrogen is a colorless, odorless, tasteless, and mostly inert diatomic gas at standard conditions, constituting 78.08% by volume of Earth's atmosphere...
in the peptide bond
Peptide bond
This article is about the peptide link found within biological molecules, such as proteins. A similar article for synthetic molecules is being created...
. If the protein is folded, the peaks are usually well dispersed, and most of the individual peaks can be distinguished. Being a relatively cheap and quick experiment, the HSQC is useful to screen candidates for structure
Protein structure
Proteins are an important class of biological macromolecules present in all organisms. Proteins are polymers of amino acids. Classified by their physical size, proteins are nanoparticles . Each protein polymer – also known as a polypeptide – consists of a sequence formed from 20 possible L-α-amino...
determination by NMR. The number of peaks in the spectrum should match the number of residues in the protein (though sidechains with nitrogen-bound protons will add additional peaks). It will probably be difficult to solve the structure of the protein if this is not the case. The labour-intensive process of structure determination is usually not undertaken until a good HSQC is obtained.
It is not possible to assign the HSQC spectrum by itself, or in other words to determine which peaks correspond to a particular residue in the protein. This process can be done in different ways as outlined in the protein NMR article. The assignment of the spectrum is usually the first step in a structure determination, and is essential for a meaningful interpretation of more advanced NMR experiments.
The HSQC experiment is also useful for detecting interactions with ligand
Ligand
In coordination chemistry, a ligand is an ion or molecule that binds to a central metal atom to form a coordination complex. The bonding between metal and ligand generally involves formal donation of one or more of the ligand's electron pairs. The nature of metal-ligand bonding can range from...
s, such as other proteins or drugs. By comparing the HSQC of the free protein with the one bound to the ligand, it is possible to find the changes in the chemical shift
Chemical shift
In nuclear magnetic resonance spectroscopy, the chemical shift is the resonant frequency of a nucleus relative to a standard. Often the position and number of chemical shifts are diagnostic of the structure of a molecule...
s of the peaks, which is most likely to occur in the binding interface.
See also
- Protein NMR
- Nuclear Magnetic ResonanceNuclear magnetic resonanceNuclear magnetic resonance is a physical phenomenon in which magnetic nuclei in a magnetic field absorb and re-emit electromagnetic radiation...
- Protein structureProtein structureProteins are an important class of biological macromolecules present in all organisms. Proteins are polymers of amino acids. Classified by their physical size, proteins are nanoparticles . Each protein polymer – also known as a polypeptide – consists of a sequence formed from 20 possible L-α-amino...
- Structural biologyStructural biologyStructural biology is a branch of molecular biology, biochemistry, and biophysics concerned with the molecular structure of biological macromolecules, especially proteins and nucleic acids, how they acquire the structures they have, and how alterations in their structures affect their function...
General references
- Protein NMR Spectroscopy : Principles and Practice (1995) John Cavanagh, Wayne J. Fairbrother, Arthur G. Palmer III, Nicholas J. Skelton, Academic Press
External links
- Protein NMR Protein NMR spectra