Folding intermediate states

It has been shown that under standard conditions most singledomain globular proteins exhibit a folding/unfolding behavior consistent with the two-state mechanism (Freire and Biltonen, 1978a Privalov, 1979). From a statistical thermodynamic standpoint the implication is that the population of partially folded intermediate states is negligible and the partition function reduces to two terms ... 

Florida Memorial College, 1, 212 fluctuation theorem, 1,109 fluid properties, 1, 239-244 focal-point approach (FPA), 1, 39 3,160 folding intermediate states, 3, 9 force fields, 3,162... 

K. The fact that different experimental probes (42 NH sites and one scence probe) exhibit distinct kinetic behavior demonstrates that partially folded intermediate states are populated on the folding pathway of cyt c. 

Aggregation. The existence of 1 (folding intermediate) states in several proteins leads to the inactivation of the protein by aggregation. Moderate amounts of stress can generate 1 states, which retains secondary structure but tertiary structural... 

A study of the tetrameric transthyretin (TTR) complex by means of TWIM-MS, computational modeling, and activation of selected ions by acceleration in the source region of the instrument, revealed that during the dissociation of protein complexes a number of partially-folded intermediate states were stable during the time-scale of the experiment (5-25 ms) [75], The results indicated that upon activation, unfolding of one or more subunits occurred while the remainder of the subunits remained folded. This occasion was the first time that partially-folded intermediates had been studied such types of measurements will allow future studies into the understanding of the dissociation mechanism of other large protein complexes to be carried out. 

A number of proteins are known to pass through a transient intermediate state, the so-called molten globule state. The precise stmctural features of this state are not known, but appear to be compact, and contain most of the regular stmcture of the folded protein, yet have a large side-chain disorder (9). 

Recently Alan Fersht, Cambridge University, has developed a protein engineering procedure for such studies. The technique is based on investigation of the effects on the energetics of folding of single-site mutations in a protein of known structure. For example, if minimal mutations such as Ala to Gly in the solvent-exposed face of an a helix, destabilize both an intermediate state and the native state, as well as the transition state between them, it is likely that the helix is already fully formed in the intermediate state. If on the other hand the mutations destabilize the native state but do not affect the energy of the intermediate or transition states at all, it is likely that the helix is not formed until after the transition state. 

In the native protein these less stable ds-proline peptides are stabilized by the tertiary structure but in the unfolded state these constraints are relaxed and there is an equilibrium between ds- and trans-isomers at each peptide bond. When the protein is refolded a substantial fraction of the molecules have one or more proline-peptide bonds in the incorrect form and the greater the number of proline residues the greater the fraction of such molecules. Cis-trans isomerization of proline peptides is intrinsically a slow process and in vitro it is frequently the rate-limiting step in folding for those molecules that have been trapped in a folding intermediate with the wrong isomer. 

Implicit in the presumption of folding pathways is the existence of intermediate, partially folded conformational states. The notion of intermediate states on the pathway to a tertiary structure raises the possibility that segments of a protein might independently adopt local and well-defined secondary structures (a-helices and /3-sheets). The tendency of a peptide segment to prefer a particular secondary structure depends in turn on its amino acid composition and sequence. 

ISF). However, in the P6s crystal form ( intermediate state ), the cluster binding subdomain appears to be detached from the base fold fi sheets 1 and 2) in an open conformation 42). If the functional domains of the two crystal forms are superposed using the base fold residues, the rms deviation of the Ca positions of the cluster binding... 

REX-CPHMD simulations have also been applied to understand the mechanism of the formation of protein intermediate states. Recent solution NMR data revealed a sparsely populated intermediate in the villin headpiece domain, in which the N-terminal subdomain is largely random but the C-terminal subdomain adopts a nativelike fold [34], Interestingly, H41 in this intermediate state titrates at a pH value of... 

Fink, A. L. (1995). Compact intermediate states in protein folding. Annu. Rev. Biophys. Biomol. Struct. 24, 495-522. 

---