Protein Unstructured

Proteins in solution are typically more mobile than those in a crystal. At equilibrium the protein in solution will fluctuate and its conformation is best represented as an ensemble of structures. This is not so bad The flexibility can leave certain portions of the protein unstructured. The mobile portions of the protein are often functionally important. Therefore, one can appreciate the disorder in the refined solution phase structure as a clue to the function of the protein and its equilibrium structure. When we seek to determine a protein s structure at room temperature we must appreciate that the native state consists of an ensemble of structures. 

They started from the sequence of a domain, Bl, from an IgG-binding protein called Protein G. This domain of 56 amino acid residues folds into a four-stranded p sheet and one a helix (Figure 17.16). Their aim was to convert this structure into an all a-helical structure similar to that of Rop (see Chapter 3). Each subunit of Rop is 63 amino acids long and folds into two a helices connected by a short loop. The last seven residues are unstructured and were not considered in the design procedure. Two subunits of Rop form a four-helix bundle (Figure 17.16). 

Caseins, however, do not display a distinctive conformational transition upon denaturation and texturization they are considered a family of natively unstructured proteins (Farrell et ah, 2006a,b). 

This chapter has reviewed the application of ROA to studies of unfolded proteins, an area of much current interest central to fundamental protein science and also to practical problems in areas as diverse as medicine and food science. Because the many discrete structure-sensitive bands present in protein ROA spectra, the technique provides a fresh perspective on the structure and behavior of unfolded proteins, and of unfolded sequences in proteins such as A-gliadin and prions which contain distinct structured and unstructured domains. It also provides new insight into the complexity of order in molten globule and reduced protein states, and of the more mobile sequences in fully folded proteins such as /1-lactoglobulin. With the promise of commercial ROA instruments becoming available in the near future, ROA should find many applications in protein science. Since many gene sequences code for natively unfolded proteins in addition to those coding for proteins with well-defined tertiary folds, both of which are equally accessible to ROA studies, ROA should find wide application in structural proteomics. 

Fig. 10. Comparison of VCD spectra of four proteins in H2O (left, amide I + II) and D2O (right, amide V + IF) with dominant secondary structure contributions from G -helix (myoglobin, MYO, top), /3-sheet (immunoglobin, IMUN), both helix and sheet (lactoferrin, LCF) and no structure (o -casein, CAS, bottom). The comparisons emphasize the distinct band shapes developed in the amide I and V for each structural type. Note the reduced S/N in the F O-based spectra and the shape changes upon H/D exchange for helix and sheet (and mixed) structures, but relatively little for the unstructured CAS.

Thermally denatured proteins have been studied for a variety of systems using FTIR and VCD. The resulting high-temperature spectra often reflect the characteristics seen earlier for random coil peptides as well as that seen for the unstructured casein. Particularly the amide I IR bands show a frequency shift to center on a broadened band at 1645-50 cm-1. The amide I VCD loses its distinctive character (Fig. 11) and tends toward... 

Recently, there has been great interest in proteins that exhibit biological activity but lack a well-defined secondary or tertiary structure after purification (Dunker et al., 1998, 2001 Schweers et al., 1994 Uversky et al., 2000 Wright and Dyson, 1999). Such proteins are referred to as intrinsically disordered or unstructured. An analysis in 1998 of the Swiss Protein Database revealed that about 15,000 proteins in that database are likely to contain disordered segments at least 40 residues in length (Romero et al., 1998). Dyson and Wright (2002) review intrinsically disordered proteins in this volume. 

B. Characterization of Proteins That Are Unstructured under Nondenaturing Conditions... 

C. Intrinsically Unstructured Proteins Coupled Folding and Binding Events... 

There are now numerous examples of proteins that are unstructured or only partially structured under physiological conditions yet are nevertheless functional (Dunker and Obradovic, 2001 Wright and Dyson, 1999). In many cases, such intrinsically disordered proteins adopt folded structures upon binding to their biological targets. As the proteins that constitute the transcriptional machinery have become... 

These results strongly suggest that unstructured peptides have definite backbone conformations and that the concept of a denatured protein as a structureless random chain breaks down when backbone conformations of individual residues are described, although the random chain concept may still be useful when describing the overall chain conformation. 

Ohashi, T., Galiacy, S. D., Briscoe, G. and Erickson, H. P. (2007). An experimental study of GFP-based FRET, with application to intrinsically unstructured proteins. Protein Sci. 16, 1429-38. 

Tompa, P. (2002). Intrinsically unstructured proteins. Trends Biochem. Sci. 27, 527-533. Tompa, P. (2005). The interplay between structure and function in intrinsically unstructured proteins. FEBS Lett. 579, 3346-3354. 

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