Differential rate constants of racemization of aspartyl and asparaginyl residues in human alpha A-crystallin mutants.

Asp58 and Asp151 in alpha A-crystallin of human eye lenses become highly inverted and isomerized to d-beta-Asp residues with age. Racemization was previously shown to proceed rapidly when the residue on the carboxyl side of the Asp residue is small. Asn was also demonstrated to be more susceptible to racemization than Asp in protein.

Amyloid nucleation triggered by agitation of beta2-microglobulin under acidic and neutral pH conditions.

Amyloid nucleation through agitation was studied with beta2-microglobulin, which is responsible for dialysis-related amyloidosis, in the presence of salt under acid and neutral pH conditions. First, the aggregation of beta2-microglobulin in NaCl solutions was achieved by mildly agitating for 24 h at 37 degrees C protein solutions in three different states: acid-unfolded, salt-induced protofibrillar, and native. The formation of aggregates was confirmed by an increase in light scattering intensity of the solutions.

AlphaB-crystallin, a small heat-shock protein, prevents the amyloid fibril growth of an amyloid beta-peptide and beta2-microglobulin.

AlphaB-crystallin, a small heat-shock protein, exhibits molecular chaperone activity. We have studied the effect of alphaB-crystallin on the fibril growth of the Abeta (amyloid beta)-peptides Abeta-(1-40) and Abeta-(1-42). alphaB-crystallin, but not BSA or hen egg-white lysozyme, prevented the fibril growth of Abeta-(1-40), as revealed by thioflavin T binding, total internal reflection fluorescence microscopy and CD spectroscopy.

Investigation of the protein pre-crystallization solution using analytical ultracentrifugation.

Analytical ultracentrifugation was used to study the crystal growth units in hen egg-white lysozyme pre-crystallization solution. Solutions containing various concentrations of lysozyme and NaCl in 50 mM sodium acetate buffer were used for experiments. The crystallization solution was ultracentrifuged using a mode where the sedimentation and diffusion are in equilibrium.

Multidrug transporter MexB of Pseudomonas aeruginosa: overexpression, purification, and initial structural characterization.

Structural and functional characterization of the multidrug transporter, MexB, of Pseudomonas aeruginosa is significantly restricted due to a low yield of approximately 0.1 mg/L of culture from natural sources. To facilitate structural studies of this medically important transporter protein, we developed a large-scale system for expression of the genetically engineered recombinant, MexB, in the Escherichia coli cell.

Metal ion-dependent effects of clioquinol on the fibril growth of an amyloid {beta} peptide.

Although metal ions such as Cu(2+), Zn(2+), and Fe(3+) are implicated to play a key role in Alzheimer disease, their role is rather complex, and comprehensive understanding is not yet obtained. We show that Cu(2+) and Zn(2+) but not Fe(3+) renders the amyloid beta peptide, Abeta(1-40), nonfibrillogenic in nature. However, preformed fibrils of Abeta(1-40) were stable when treated with these metal ions.

Critical balance of electrostatic and hydrophobic interactions is required for beta 2-microglobulin amyloid fibril growth and stability.

Investigation of factors that modulate amyloid formation of proteins is important to understand and mitigate amyloid-related diseases. To understand the role of electrostatic interactions and the effect of ionic cosolutes, especially anions, on amyloid formation, we have investigated the effect of salts such as NaCl, NaI, NaClO(4), and Na(2)SO(4) on the amyloid fibril growth of beta(2)-microglobulin, the protein involved in dialysis-related amyloidosis. Under acidic conditions, these salts exhibit characteristic optimal concentrations where the fibril growth is favored.

Seeding-dependent maturation of beta2-microglobulin amyloid fibrils at neutral pH.

Beta2-microglobulin (beta2-m) is a major component of amyloid fibrils deposited in patients with dialysis-related amyloidosis. Recent studies have focused on the mechanism by which amyloid fibrils are formed under physiological conditions, which had been difficult to reproduce quantitatively. Yamamoto et al.