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General Approach to Assignments

To illustrate the assignment process consider the following tetrapeptide sequence that is part of a protein  [Pg.53]

The assignment strategies nnder different experimental conditions (molecular weight, isotopic labeling) are discussed below. [Pg.54]

As the molecular weight increases, the sensitivity of the H TOCSY decreases owing to the loss of magnetization by efficient spin-spin relaxation. Once the molecular weight exceeds 12 to 15 kDa, the H TOCSY will provide incomplete information because many of the peaks are of weak intensity and are buried in the noise. [Pg.54]

The TOCSY experiments provide the chemical shifts of the aliphatic protons. Depending on the residue type, it is usually possible to assign a spin system to a class of amino acids. In favorable cases, it may be possible to uniquely identify the residue type of a spin system. For example, residues like Asn, Asp, Glu, and Gin are easily distinguished from hydrophobic residues based on characteristic proton chemical shifts. A number of residues, such as Ser, Ala, and Thr, can be uniquely assigned from their characteristic proton shifts. Note that if any of these residues [Pg.54]

The next step in the assignment process is to assign each spin system to a particular residue in the protein. This can be accomplished in one of two ways. The first method utilizes dipolar coupling between protons on adjacent amino acids (observed in a NOESY spectra). In a protein, one would expect that some of the protons on one amino acid are within 5 A of the protons on the adjacent amino acid. In this particular example, dipolar coupling between adjacent residues would be used to infer that Valio is adjacent to Alan, that Alan is adjacent to Ser, and so on. The difficulty with this method is that a proton will show dipolar coupling to a large [Pg.55]


Figure A2.1 General approach to assignment of handedness to a molecule with a chiral centre. In this example the groups are assumed to have priority a > b > c > d. Figure A2.1 General approach to assignment of handedness to a molecule with a chiral centre. In this example the groups are assumed to have priority a > b > c > d.

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