HNCOCA Experiment
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HNCOCA Experiment
HNCOCA is a 3D triple-resonance NMR experiment commonly used in the field of protein NMR. The name derives from the experiment's magnetization transfer pathway: The magnetization of the amide proton of an amino acid residue is transferred to the amide nitrogen, and then to the alpha carbon of the previous residue in the protein's amino acid sequence. In contrast, the complementary HNCA experiment transfers magnetization to the alpha carbons of both the starting residue and the previous residue in the sequence. The HNCOCA experiment is used, often in tandem with HNCA, to assign alpha carbon resonance signals to specific residues in the protein. This experiment requires a purified sample of protein prepared with 13C and 15N isotopic labelling, at a concentration greater than 0.1 mM, and is thus generally only applied to recombinant proteins Recombinant DNA (rDNA) molecules are DNA molecules formed by laboratory methods of genetic recombination (such as molecular cloning) tha ...
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Triple-resonance Nuclear Magnetic Resonance Spectroscopy
Triple resonance experiments are a set of multi-dimensional nuclear magnetic resonance spectroscopy (NMR) experiments that link three types of atomic nuclei, most typically consisting of 1H, 15N and 13C. These experiments are often used to assign specific resonance signals to specific atoms in an isotopically-enriched protein. The technique was first described in papers by Ad Bax, Mitsuhiko Ikura and Lewis Kay in 1990, and further experiments were then added to the suite of experiments. Many of these experiments have since become the standard set of experiments used for sequential assignment of NMR resonances in the determination of protein structure by NMR. They are now an integral part of solution NMR study of proteins, and they may also be used in solid-state NMR. Background There are two main methods of determining protein structure on the atomic level. The first of these is by X-ray crystallography, starting in 1958 when the crystal structure of myoglobin was determined ...
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Protein Nuclear Magnetic Resonance Spectroscopy
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. The field was pioneered by Richard R. Ernst and Kurt Wüthrich at the ETH, and by Ad Bax, Marius Clore, Angela Gronenborn at the NIH, and Gerhard Wagner at Harvard University, among others. Structure determination by NMR spectroscopy usually consists of several phases, each using a separate set of highly specialized techniques. The sample is prepared, measurements are made, interpretive approaches are applied, and a structure is calculated and validated. NMR involves the quantum-mechanical properties of the central core ("nucleus") of the atom. These properties depend on the local molecular environment, and their measurement provides a map of how the atoms are linked chemically, how close they are in space, and how rapid ...
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Amino Acid
Amino acids are organic compounds that contain both amino and carboxylic acid functional groups. Although hundreds of amino acids exist in nature, by far the most important are the alpha-amino acids, which comprise proteins. Only 22 alpha amino acids appear in the genetic code. Amino acids can be classified according to the locations of the core structural functional groups, as Alpha and beta carbon, alpha- , beta- , gamma- or delta- amino acids; other categories relate to Chemical polarity, polarity, ionization, and side chain group type (aliphatic, Open-chain compound, acyclic, aromatic, containing hydroxyl or sulfur, etc.). In the form of proteins, amino acid '' residues'' form the second-largest component (water being the largest) of human muscles and other tissues. Beyond their role as residues in proteins, amino acids participate in a number of processes such as neurotransmitter transport and biosynthesis. It is thought that they played a key role in enabling life ...
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HNCA Experiment
HNCA is a 3D triple-resonance NMR experiment commonly used in the field of protein NMR. The name derives from the experiment's magnetization transfer pathway: The magnetization of the amide proton of an amino acid residue is transferred to the amide nitrogen, and then to the alpha carbons of both the starting residue and the previous residue in the protein's amino acid sequence. In contrast, the complementary HNCOCA experiment transfers magnetization only to the alpha carbon of the previous residue. The HNCA experiment is used, often in tandem with HNCOCA, to assign alpha carbon resonance signals to specific residues in the protein. This experiment requires a purified sample of protein prepared with 13C and 15N isotopic labelling, at a concentration greater than 0.1 mM, and is thus generally only applied to recombinant proteins Recombinant DNA (rDNA) molecules are DNA molecules formed by laboratory methods of genetic recombination (such as molecular cloning) that bring tog ...
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Carbon-13
Carbon-13 (13C) is a natural, stable isotope of carbon with a nucleus containing six protons and seven neutrons. As one of the environmental isotopes, it makes up about 1.1% of all natural carbon on Earth. Detection by mass spectrometry A mass spectrum of an organic compound will usually contain a small peak of one mass unit greater than the apparent molecular ion peak (M) of the whole molecule. This is known as the M+1 peak and comes from the few molecules that contain a 13C atom in place of a 12C. A molecule containing one carbon atom will be expected to have an M+1 peak of approximately 1.1% of the size of the M peak, as 1.1% of the molecules will have a 13C rather than a 12C. Similarly, a molecule containing two carbon atoms will be expected to have an M+1 peak of approximately 2.2% of the size of the M peak, as there is double the previous likelihood that any molecule will contain a 13C atom. In the above, the mathematics and chemistry have been simplified, however it can ...
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Nitrogen-15
Natural nitrogen (7N) consists of two stable isotopes: the vast majority (99.6%) of naturally occurring nitrogen is nitrogen-14, with the remainder being nitrogen-15. Fourteen radioisotopes are also known, with atomic masses ranging from 10 to 25, along with one nuclear isomer, 11mN. All of these radioisotopes are short-lived, the longest-lived being nitrogen-13 with a half-life of . All of the others have half-lives below 7.15 seconds, with most of these being below 620 milliseconds. Most of the isotopes with atomic mass numbers below 14 decay to isotopes of carbon, while most of the isotopes with masses above 15 decay to isotopes of oxygen. The shortest-lived known isotope is nitrogen-10, with a half-life of . List of isotopes , - , , style="text-align:right" , 7 , style="text-align:right" , 3 , , , p ?Decay mode shown is energetically allowed, but has not been experimentally observed to occur in this nuclide. , ? , 1−, 2− , , , - , , style="text-align:r ...
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Isotopic Labeling
Isotopic labeling (or isotopic labelling) is a technique used to track the passage of an isotope (an atom with a detectable variation in neutron count) through a reaction, metabolic pathway, or cell. The reactant is 'labeled' by replacing specific atoms by their isotope. The reactant is then allowed to undergo the reaction. The position of the isotopes in the products is measured to determine the sequence the isotopic atom followed in the reaction or the cell's metabolic pathway. The nuclides used in isotopic labeling may be stable nuclides or radionuclides. In the latter case, the labeling is called radiolabeling. In isotopic labeling, there are multiple ways to detect the presence of labeling isotopes; through their mass, vibrational mode, or radioactive decay. Mass spectrometry detects the difference in an isotope's mass, while infrared spectroscopy detects the difference in the isotope's vibrational modes. Nuclear magnetic resonance detects atoms with different gyromagnetic r ...
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Concentration
In chemistry, concentration is the abundance of a constituent divided by the total volume of a mixture. Several types of mathematical description can be distinguished: '' mass concentration'', ''molar concentration'', ''number concentration'', and ''volume concentration''. The concentration can refer to any kind of chemical mixture, but most frequently refers to solutes and solvents in solutions. The molar (amount) concentration has variants, such as normal concentration and osmotic concentration. Etymology The term concentration comes from the word concentrate, from the French , from con– + center, meaning “to put at the center”. Qualitative description Often in informal, non-technical language, concentration is described in a qualitative way, through the use of adjectives such as "dilute" for solutions of relatively low concentration and "concentrated" for solutions of relatively high concentration. To concentrate a solution, one must add more solute (for example, ...
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Recombinant DNA
Recombinant DNA (rDNA) molecules are DNA molecules formed by laboratory methods of genetic recombination (such as molecular cloning) that bring together genetic material from multiple sources, creating sequences that would not otherwise be found in the genome. Recombinant DNA is the general name for a piece of DNA that has been created by combining at least two fragments from two different sources. Recombinant DNA is possible because DNA molecules from all organisms share the same chemical structure, and differ only in the nucleotide sequence within that identical overall structure. Recombinant DNA molecules are sometimes called chimeric DNA, because they can be made of material from two different species, like the mythical chimera. R-DNA technology uses palindromic sequences and leads to the production of sticky and blunt ends. The DNA sequences used in the construction of recombinant DNA molecules can originate from any species. For example, plant DNA may be joined to bacter ...
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HNCA
HNCA is a 3D triple-resonance NMR experiment commonly used in the field of protein NMR. The name derives from the experiment's magnetization transfer pathway: The magnetization of the amide proton of an amino acid residue is transferred to the amide nitrogen, and then to the alpha carbons of both the starting residue and the previous residue in the protein's amino acid sequence. In contrast, the complementary HNCOCA experiment transfers magnetization only to the alpha carbon of the previous residue. The HNCA experiment is used, often in tandem with HNCOCA, to assign alpha carbon resonance signals to specific residues in the protein. This experiment requires a purified sample of protein prepared with 13C and 15N isotopic labelling, at a concentration greater than 0.1 mM, and is thus generally only applied to recombinant proteins. The spectrum produced by this experiment has 3 dimensions: A proton axis, a 15N axis and a 13C axis. For residue i peaks will appear at and , while for ...
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Academic Press
Academic Press (AP) is an academic book publisher founded in 1941. It was acquired by Harcourt, Brace & World in 1969. Reed Elsevier bought Harcourt in 2000, and Academic Press is now an imprint of Elsevier. Academic Press publishes reference books, serials and online products in the subject areas of: * Communications engineering * Economics * Environmental science * Finance * Food science and nutrition * Geophysics * Life sciences * Mathematics and statistics * Neuroscience * Physical sciences * Psychology Well-known products include the ''Methods in Enzymology'' series and encyclopedias such as ''The International Encyclopedia of Public Health'' and the ''Encyclopedia of Neuroscience''. See also * Akademische Verlagsgesellschaft (AVG) — the German predecessor, founded in 1906 by Leo Jolowicz (1868–1940), the father of Walter Jolowicz Walter may refer to: People * Walter (name), both a surname and a given name * Little Walter, American blues harmonica player Marion Wa ...
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