Native-like structure

In this model (Fay et al., 2005), portions of the N- and C-terminal regions, specifically residues 6 and 137, are in close proximity in the fibril, and the C-terminal domain retains a native-like structure. There is evidence that this non-amyloid-like fibril can convert to the cross-/ -containing filament with heat treatment, incubation at low pH (Bousset et al., 2003), or extensive drying (Fay et al., 2005), but it is unclear what sort of structural change might link the two fibril types. 

In a further study, Taniuchi et al. (1977) have shown that in the association of overlapping fragments of staphylococcal nuclease, two different species of active enzyme are formed. On the basis of the products of limited proteolysis, structures for the two species were deduced. In one case a structure is proposed in which fragment 1-126 assumes native-like structure over the sequence 1-48, and all of fragment 50-149 assumes native-like structure. In the other case the structure is one in which fragment 1-126 assumes native-like structure over the sequence 1-110, while that part of fragment 50-149 in the sequence interval 111-149 assumes native-like structure. The interest of these results is enhanced by the finding that the two active species initially form in relative concentrations substantially different from their equilibrium concentrations. Thus, both a mobile equilibrium and substantial kinetic control of the early products are evident. Taniuchi et al. did not reach a clear-cut mechanistic conclusion from their studies. 

Affinity chromatography was carried out on columns prepared with lightly carboxymethylated chitin, which is known to be a poor substrate for lysozyme. Both native lysozyme and regenerated 13-105 were bound to the column at pH 7 and eluted at pH 3. As controls, the basic proteins cytochrome c and pancreatic RNase A, as well as concanavalin A and a-amylase, were not bound from the same solvent at pH 7. These findings constitute a third line of evidence for formation of native-like structure in regenerated 13-105. 

The kinetics of disulfide formation, the demonstration of specific binding, and the immunochemical results all support the conclusion that native-like structure results from the oxidative folding of reduced peptide 13-105. These three independent lines of evidence support the conclusion that lysozyme has a continuous chain independent assembly region somewhere in the sequence 13-105. 

Results of several experiments in which the return of native-like structure to various parts of reduced albumin was obtained are shown in Fig. 7. It is clear that different antigenic determinants are formed... 

Fontana and Vita took the foregoing experiments as evidence not only that native-like structure persists in two fragments of thermo-lysin, but also that folding to native-like structure can occur with the denatured fragments. 

There was neither report of yield nor investigation of the kinetics of return of native characteristics. Attempts to prepare intact fragment 16-125 have not yet been successful. The immunochemical and substrate binding evidence support the claim that refolding to native-like structure did indeed occur. It clearly would be desirable to have more evidence on this system. 

Folding to native-like structure has been demonstrated with fragments of jS-galactosidase, lysozyme, serum albumin, penicillinase, and tryptophan synthetase. The capability of protein fragments for independent formation of structure therefore has substantial experimental basis. This generalization also makes plausible the idea that, in general, protein folding occurs by parts, that is, in a modular fashion. 

The stability of peptides is generally increased when natural amino acids are substituted by fluorinated analogs (e.g., tri- or hexafluoroleucine, hexafluorova-line). Such stabilization augments with the number of hexafluoroleucine residues introduced [77]. Native-like structure was preserved, but the peptides had a more structured backbone and less fluid hydrophobic core. Substitution of four leucine residues by trifluoroleucines in the leucine zipper peptide GCN4-p1d led to a substantial gain in thermal stability and resistance to chemical denaturation of the... 

H.Y. Lee, K.H. Lee, H.M. Al-Hashimi, E.N.G. Marsh, Modulating protein structure with fluorous amino acids Increased stability and native-like structure conferred on a 4-helix bundle protein by hexafluoroleucine, J. Am. Chem. Soc. 128 (2006) 337-343. 

Sherman, M.A. Chen, Y. Mas, M.T. An engineered amino-terminal domain of yeast phosphoglycerate kinase with native-like structure. Protein Sci., 6, 882-891 (1997)... 

Z-score analysis leads to a lot of result values (Figure 11.6). In general, zp-comb, zp-pair, and zp-surf together with the respective ranks (which generally should be 1 for native-like structures) are sufficient for judging the result. 

Conformationally rigid cavitands III have been used for the synthesis of four-helix bundle caviteins 29,60,61 which are de novo proteins composed of cavitands and proteins (Figure 4). The proximity of four a-helical peptides significantly stabilizes their native-like structure, which was proved by denaturation experiments with guanidine hydrochloride.62... 

Sambashivan S, Liu Y, Sawaya MR, Gingery M, Eisenberg D. Amyloid-like fibrils of ribonuclease A with three-dimensional domain-swapped and native-like structure. Nature 2005 437 266-269. 

Simulation-derived key active site structural parameters are provided in Table 4, and representative hydrogen-bond base pairing at the C3-G8 positions are shown in Figure 5. In addition, a set of control simulations were performed on a benign U7C mutation, and the wild-type simulation with the active site Mg2+ ion removed. An implicit assumption herein is that the mutated sequences fold to a native-like structure. 

Likewise, the strucmre of subtilisin (pH 3.0) suspended in varying ratios of acetonitrile and water demonstrated a-helical content similar to that in the lyophilized powder (Griebenow and Klibanov, 1996). Furthermore, the rate of transesteriflcation reactions of subtilisin (pH 7.8) suspended in DMSO/acetonitrile, formamide/acetonitrile or formamide/dioxane were increased approximately 100-fold over aqueous conditions (Almarsson and Klibanov, 1996). Similar results were obtained for subtilisin (pH 7.8) in a tetrahydrofuran/1-propanol mixture (Affleck et al., 1992). These results can be attributed to the increased structural rigidity of the active conformation of the protein in the solid, and the denaturing characteristics of the solvent at the solvent-particulate interface. Preservation of this molecular memory or molecular imprint of the protein can also be used to stabilize structure and activity (Mishra et al., 1996 Rich and Dordick, 1997 Santos et al., 2001). Subtilisin was lyophilized from crown ethers, resulting in more native like structure, by FTIR, and increased enzyme activity in THF, acetonitrile and dioxane (Santos et al.,2001). 

Lau and Dill have also investigated the statistical mechanics of folding for simplified protein models on two-dimensional square lattices. They explored both conformational space (the set of all possible conformations) and sequence space (the set of all possible sequences) and concluded that many sequences have stable, compact, native-like structures. Another conclusion of these studies was that sequences tended to form a single, unique structure, even with only two types of residues (hydrophobic and polar). This tendency increased with chain length. Moreover, one or two mutations in these sequences did not greatly destabilize most folded states. 

---