Globular proteins using coiled coils

Coiled coils are frequently used to form oligomers of fibrous and globular proteins... 

The leucine zipper DNA-binding proteins, described in Chapter 10, are examples of globular proteins that use coiled coils to form both homo- and heterodimers. A variety of fibrous proteins also have heptad repeats in their sequences and use coiled coils to form oligomers, mainly dimers and trimers. Among these are myosin, fibrinogen, actin cross-linking proteins such as spectrin and dystrophin as well as the intermediate filament proteins keratin, vimentin, desmin, and neurofilament proteins. 

Early attempts used data obtained from homopolypeptides, such as poly(Lys), for their basis spectra [87, 88]. In the past fifteen years, approaches using data from globular proteins have emerged [18, 89-101]. Basically, a data base comprised of proteins with known secondary structure compositions is assembled and far UV CD spectra recorded. The choice of the proteins to be included is critical and various combinations have been examined. Mathematical matrix methods can be used to extract basis spectra which represent the contributions from the various secondary structures. Typically, four or five basis spectra can be obtained (corresponding to a helices, jj sheets, p turns, and random coil structures). In some approaches, such as those developed by Johnson and co-workers [11, 12, 51, 52, 102], separate basis spectra can be obtained for parallel and antiparallel p sheets. These basis spectra are then linearly combined to reconstruct the CD spectrum of the protein of interest The proportion of the basis spectra used to provide the best fit to the spectrum corresponds to the percentage of that secondary structure in the protein of interest Complete details of the mathematical algorithms that have been employed can be found elsewhere [10, 12, 17, 89, 103]. 

When using molar mass as the calibration parameter, one has to consider the shape of macromolecules in solution during the data evaluation e.g., it is not possible to calculate directly the molar mass of the randomly coiled proteins from the calibration curve obtained by means of the globular proteins. 

One of the most useful ways of using CD spectroscopy is to deduce the relative proportions of secondary structure elements in a globular or even in a fibrous protein. The main structural elements in a globular protein that can be identified separately by CD spectroscopy are defined as a-helix, /1-sheets or random coil. Each main secondary structural element is right handed... 

EXAMPLE 4.8 Many globular proteins contain elements of a helix, P structures, plus random-coil regions. It is instructive to draw these structures by using the stylized forms shown in Fig. 4-12, in which a helices are represented... 

It is cautioned, however, that the use of such a database from solution NMR is not always appropriate for the interpretation of solid-state NMR data, because of a possible source of errors arising from the chemical shift references for the solid and solution NMR studies. Moreover, further larger dispersions of chemical shifts from the solution NMR of globular proteins cannot always be ignored. Such data, however, are not always free from ambiguities in choosing the appropriate chemical shifts of the random coil, which are essential for data collection in solution NMR. These are simply treated as hidden parameters, and not always as necessary parameters for the interpretation of the solid-state NMR, unless NMR data of the fully hydrated samples are concerned. In fact, the chemical shift data available from such databases are in many instances biased as a reference for solid-state NMR because of choice of reference data for both solid and solution NMR. It is advised therefore to utilize the database from the solid state, if one aims to interpret the chemical shift data from solid-state NMR. " ... 

Next, we shall review a method of constructing a "universal calibration curve" that has been used extensively by high-polymer chemists and may deserve closer attention from biopolymer chemists as well. Second, we will review our own efforts to relate elution volumes obtained in SEC to the hydrodynamic and statistical parameters of random coil polymers and globular proteins. 

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