Resonance assignments general approach

There are two general approaches through which stage (1), resonance assignment, can be accomplished, and the choice between them is determined essentially by the molecular mass of the protein. The first approach, which was pioneered in the laboratory of Kurt Wiithrich34 and which led to his award of the 2002 Nobel Prize in Chemistry, involves the use of 2D 1H—1 H (homonuclear) NMR experiments such as COSY, NOESY, and TOCSY (see below). This approach is still widely used today, but only for proteins smaller than lOkDa that cannot be isotopically labeled. Nevertheless, the homonuclear NMR strategy is suitable for studying peptides purified from natural sources, and this will be the focus of Section 9.09.3. 

Since there are six observable tetrads and ten observable pentads, it is a formidable task to make resonance assignments for these structxiral features. However, there are a number of general approaches for doing this. These include (a) the use of stationary or "necessary n-add relationships" to correlate the relative intensities of resonances (b) the development of empirical relationships to correlate differences in chemical shift due to structural differences ... 

Sometimes more than two empirical rules must be developed to make proper assignments, but it is seldom necessary to use more than three or four rules. This general approach can also be used for assigning resonances due to stereosequences in homopolymers, as has been mentioned in the discussion on stereoregular homopolymers. Klesper employed four rules to completely assign the a-methyl proton resonances of atactic meth-acrylic acid-methyl methacrylate copolymers [27]. Since he was the first to develop and apply rules of the sort considered here, it seems appropriate to refer to them in a general sense as "Klesper s rules."... 

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