Molten globules native state

Fig. 33. Schematic representation of the effects of pressure on oligomeric proteins a) native dimeric protein with cavities/voids b) dissociation of the oligomer, hydration with electrostriction of polar/ionic groups, hydrophobic hydration of unpolar groups (-CR), release of void volume c) weakening of hydrophobic interactions provides pathways for water to penetrate into the interior of the protein, swelling of the core - molten-globule like state d) unfolding of subunits, disruption of the secondary/tertiary structure (hydration of residues not plotted here), loss of cavity volume within protein (adopted from ref. 139).

Figure 6.2 The molten globule state is an important intermediate in the folding pathway when a polypeptide chain converts from an unfolded to a folded state. The molten globule has most of the secondary structure of the native state but it is less compact and the proper packing interactions in the interior of the protein have not been formed.

The collapse of the unfolded state to generate the molten globule embodies the main mystery of protein folding. What is the driving force behind the choice of native tertiary fold from a randomly oriented polypeptide chain ... 

The conformational plasticity supported by mobile regions within native proteins, partially denatured protein states such as molten globules, and natively unfolded proteins underlies many of the conformational (protein misfolding) diseases (Carrell and Lomas, 1997 Dobson et al., 2001). Many of these diseases involve amyloid fibril formation, as in amyloidosis from mutant human lysozymes, neurodegenerative diseases such as Parkinson s and Alzheimer s due to the hbrillogenic propensities of a -synuclein and tau, and the prion encephalopathies such as scrapie, BSE, and new variant Creutzfeldt-Jacob disease (CJD) where amyloid fibril formation is triggered by exposure to the amyloid form of the prion protein. In addition, aggregation of serine protease inhibitors such as a j-antitrypsin is responsible for diseases such as emphysema and cirrhosis. 

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. 

The principal defining properties of the molten globule are as follows (Arai and Kuwajima, 2000) (1) substantial secondary structure (2) no significant tertiary structure (3) structure only slightly expanded from the native state (10—30% increase in radius of gyration) (4) a loosely packed hydrophobic core with increased solvent accessibility. The first two criteria are readily assessed by far- and near-UV CD, respectively. Therefore, CD has been extensively applied to the detection and characterization of molten globules. 

Carbonic anhydrase is another protein that forms a compact A-state at low pH (Wong and Hamlin, 1974). In this case, the far-UV CD changes on going from native protein to molten globule are quite spectacular, as illustrated in Figure 38. At neutral pH the protein has a rather weak... 

A puzzling problem was posed by Levinthal many years ago.329 We usually assume that the peptide chain folds into one of the most stable conformations possible. However, proteins fold very rapidly. Even today, no computer would be able, in our lifetime, to find by systematic examination the thermodynamically most stable conformation.328 It would likewise be impossible for a folding protein to "try out" more than a tiny fraction of all possible conformations. Yet folded and unfolded proteins often appear to be in a thermodynamic equilibrium Experimental results indicate that denatured proteins are frequently in equilibrium with a compact denatured state or "molten globule" in which hydrophobic groups have become clustered and some secondary structures exists.330-336 From this state the polypeptide may rearrange more slowly through other folding intermediates to the final "native conformation."3363 3361 ... 

Fig. 7.6. Figure 7.6. Backscattered ICP Raman (IR f IL) ancj j oA (IR - IL) spectra of (a) human lysozyme in the native state, (b) human lysozyme in the low pH molten globule state, and (c) the T-A-l peptide from wheat glutenin. Adapted from references 45 and 46...

Figure 36 shows the femtosecond-resolved fluorescence transients of A144W mutant in the native (pH = 6.0) and molten globule (pH = 4.0) states [198] for... 

Figure 36. Fs-resolved fluorescence transients of mutant A144W for several gated emission wavelengths in the native and molten globule states in short (left) and long (right) time ranges. Note that all of the signals become faster in the molten globule state.

Figure 38. Hydration correlation functions c(t) of 16 mutants in both native (N, circles) and molten globule (MG, squares) states. The solid lines are the best biexponential fit to c t). The insets show the local protein environment around sites of mutation both in surface map and ribbon representation. On surface maps, white, light gray, and dark gray colors represent nonpolar, positively, and negatively charged residues, respectively, and mutation sites are shown in black. On ribbon structures, mutation sites are indicated with black balls, and the A-H letters indicate identities of local helices.

---