Human serum albumin (HSA) is known to exist as N (pH approximately 7), B (pH approximately 9), and F (pH approximately 3.5) isomeric forms and an equilibrium intermediate state (I) accumulate in the urea induced unfolding pathway of HSA around 4.8-5.2 M urea concentrations. These states displayed characteristic structure and functions. To elucidate the ciprofloxacin (CFX) binding behavior of HSA, the binding of ciprofloxacin with these conformational states of human serum albumin (HSA) has been investigated by fluorescence spectroscopy. The binding constant (K) for N, B, F, and I conformation of HSA were 6.92 x 10(5), 3.87 x 10(5), 4.06 x 10(5), and 2.7 x 10(5) M(-1) and the number of binding sites (n) were 1.26,1.21, 1.16, and 1.19, respectively. The standard free energy changes (DeltaGbinding(0)) of interaction were found to be -33.3 (N isomer), -31.8 (B isomer), -32 (F isomer), and -30.0 kJ mol(-1) respectively. By using unfolding pathway of HSA, domain II of HSA has been assigned to possess binding site of ciprofloxacin. Plausible correlation between stability of CFX-N and CFX-B complexes and drug distribution have been discussed. At plasma concentration of HSA fraction of free CFX, which contributes potential to its rate of transport across cell membrane, was found to be approximately 80% more for B isomers compared to N isomers of HSA. The conformational changes in two physiologically important isomers of HSA (N and B isomers) upon ciprofloxacin binding were evaluated by measuring far, near-UV CD, and fluorescence properties of the CFX-HSA complex.

Background: ␣-Synuclein is an aggregation-prone protein that reconfigures more slowly under aggregating conditions. Results: Curcumin binds to monomeric ␣-synuclein, prevents aggregation, and increases the reconfiguration rate, particularly at high temperatures. Conclusion: Curcumin rescues the protein from aggregation by making the protein more diffusive. Significance: The search for aggregation inhibitors should account for changes in chain dynamics by the small molecule.

We hypothesize that the first step of aggregation of disordered proteins, such as α-synuclein, is controlled by the rate of backbone reconfiguration. When reconfiguration is fast, bimolecular association is not stable, but as reconfiguration slows, association is more stable and subsequent aggregation is faster. To investigate this hypothesis, we have measured the rate of intramolecular diffusion in α-synuclein, a protein involved in Parkinson's disease, under solvent conditions that accelerate or decelerate aggregation. Using the method of tryptophan-cysteine (Trp-Cys) quenching, the rate of intramolecular contact is measured in four different loops along the chain length. This intrinsically disordered protein is highly diffusive at low temperature at neutral pH, when aggregation is slow, and compacts and diffuses more slowly at high temperature or low pH, when aggregation is rapid. Diffusion also slows with the disease mutation A30P. This work provides unique insights into the earliest steps of α-synuclein aggregation pathway and should provide the basis for the development of drugs that can prevent aggregation at the initial stage.unfolded state | Alzheimer's disease | amyloid T he protein α-synuclein is the primary component of intracellular aggregants known as Lewy bodies, a hallmark of Parkinson's disease, and likely plays a central role in neuronal cell death (1). When recombinantly expressed and purified it is intrinsically disordered in solution and highly prone to aggregation (2). Recent work has shown that the native structure in human cells is likely a helical tetramer that is resistant to aggregation, but when this tetramer is denatured the monomers cannot autonomously refold and aggregate very quickly (3, 4). Thus it is likely that the aggregation pathway passes through the monomer. In vitro, the aggregation propensity of α-synuclein is very sensitive to solution conditions such as temperature, pH, salt concentration, and solution cofactors. There has been significant work to discover the structures that form in association with lipids, structures of final aggregates, and structures that may be the aggregation precursor (5-15). However, the underlying reason for why monomeric α-synuclein is so prone to aggregation is still unknown. We propose that a physical influence on aggregation is the internal dynamics of the disordered chain, that is, the rate of intramolecular diffusion.Over the past decade, a number of groups have measured intramolecular diffusion in a variety of sequences using measurements of intramolecular contact between two probes placed in the chain. All unstructured peptides, including fragments of α-synuclein, diffuse relatively rapidly, approximately 1-3 × 10 −6 cm 2 s −1 (16-18). These rates are less than 10 times slower than free diffusion of individual amino acids and about the same as translational diffusion of the entire chain, suggesting that the chain does not exert significant drag on the movement of individual segments (19), although there are measurable tail effects when ...

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