Characterization of oxidation end product of plasma albumin 'in vivo'.

Anti-oxidants are paradoxically much lower in plasma than inside cells even blood is comparably exposed to the oxidative stress. 'In vitro' models suggest a critical role of albumin as substitutive anti-oxidant in plasma but no proof for this role is available 'in vivo.' Herein, we demonstrate by LC/MS/MS that plasma albumin undergoes massive oxidation in primary nephrotic syndrome, involving stable sulphonation SO3- of the free SH of Cys 34 with +48Da increase in exact mass of the protein (ESI-MS) and formation of a fast moving isoform in the pH range between 5 and 7.

Thermodynamics of proteins in unusual environments.

Some aspects of protein thermodynamics in unconventional environments are addressed and discussed. Aqueous medium, especially dilute solution is the 'usual' ambient, which mediates all the interactions between protein and nearby molecules. When the water content is low, the surroundings may be considered 'unusual', exerting new stresses on the protein molecule and demanding different responses and property changes.

Analytical titration curves of glycosyl hydrolase Cel45 by combined isoelectric focusing-electrophoresis.

The electrical charge of endocellulase Cel45-core has been determined by combined isoelectric focusing-electrophoresis in the range of pH 3-9. In order to transform electrophoretic mobility to absolute electrical charge value, several corrections were applied: the frictional coefficient theoretically calculated from the molecular dimensions depends on porous gel structure and on the ionic strength of the solution. By comparing the curve calculated according to the Linderstrom-Lang equation, the number of charged electrical groups exposed to the solvent and their apparent ionization constants, pK(o)i, can be determined.

Decoupling of melting domains in immobilized ribonuclease A.

Ribonuclease A has been immobilized on silica beads through glutaraldeyde-mediated chemical coupling in order to improve the stability of the protein against thermal denaturation. The thermodynamic and binding properties of the immobilized enzyme have been studied and compared with those of the free enzyme. The parameters describing the binding of the inhibitor 3'-CMP (Ka and delta H) as monitored by spectrophotometry and calorimetry were not significantly affected after immobilization.

Differential scanning calorimetry characterization of rabbit brain membrane fractions.

Various membrane fractions have been prepared from rabbit brain synaptosomes by centrifugation on discontinuous sucrose gradients after osmotic shock. These fractions were characterized by electron microscopy (E.M.