'Smart' polymers and what they could do in biotechnology and medicine.

Stimulus-responsive or 'smart' polymers undergo strong conformational changes when only small changes in the environment (e. g. pH, temperature, ionic strength) occur. These changes result in phase separation from aqueous solution or order-of-magnitude changes in hydrogel size. Smart polymers are used in bioseparation and drug delivery, for the development of new biocatalysts, as biomimetic actuators, and as surfaces with switchable hydrophobic-hydrophilic properties.     

Activity and properties of L-lysine amidase in Salmonella strains

Distribution, activity level and properties of alpha-lysinamidase have been studied in Salmonella strains. The Km value for L-lysinamide was calculated to be 4.2 mM and for L-alpha-aminocaprolactame--5.1 mM. This enzyme, parallel with lysinamide, catalyzes hydrolysis of alpha-aminocaprolactam and leucinamide. Asparagine, glutamine, caprolactam, triptophanamide were not lysinamidase substrates.     

Affinity precipitation of monoclonal antibodies by nonstoichiometric polyelectrolyte complexes

The nonstoichiometric polyelectrolyte complex (PEC) formed by poly(methacrylic acid) (degree of polymerization 1830) (PMAA)and poly(N-ethyl-4-vinyl-pyridinium bromide) (degree of polymerization 530) (PEVP) undergoes reversible precipitation from aqueous solution at any desired pH-value in the range 4.5–6.5 depending on the ionic strength and PEVP/PMAA ratio in the complex. The antigen, inactivated glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from rabbit was covalently coupled to PEVP. The resulting GAPDH–PEVP/PMAA complex was used for the purification of antibodies from a 6G7 clone specific towards inactivated GAPDH. The crude extract was incubated with GAPDH-containing PEC and the precipitation of the PEC was carried out at 0.01 M NaCl and pH 4.5, 5.3, 6.0 and 6.5 using PEC with PEVP/PMAA ratios of 0.45, 0.3, 0.2 and 0.15, respectively. Purified antibodies were eluted at pH 4.0 where PECs of all compositions used were insoluble.PEC precipitation is accompanied only by small nonspecific coprecipitation of proteins. Precipitated PEC could be dissolved at pH 7.3 and used repeatedly. This revised version was published online in July 2006 with corrections to the Cover Date.     

New methods of protein purification. Affinity ultrafiltration.

This review describes a recently developed method for protein purification-affinity ultrafiltration. In affinity ultrafiltration, the protein to be purified is complexed with a macroligand composed of a soluble polymer or an insoluble microparticle with covalently bound, target protein-specific affinity ligands. The complex is trapped by an ultrafiltration membrane, whereas unwanted proteins pass through the membrane. The unwanted proteins are removed from the system by the carrier liquid. The system is then supplemented with an agent eluting the target protein by dissociating it from the microligand complex. The purified protein then passes the membrane, while the macroligand is trapped by it. The macroligand can be re-used after regeneration. Affinity ultrafiltration has a number of advantages over other protein purification techniques: 1) commercial availability of ultrafiltration systems with various high-productivity designs; 2) availability of presynthesized macroligands, which can be supplemented with additional, easily manufactured, commercial latex-based macroligands; 3) rapid separation of large solution volumes; 4) repeated use of equipment, enabling consecutive purification of different proteins; 5) simple scale-up and automation procedures.     

Aromatic amino acid metabolism in Enterobacteriaceae microorganisms]

The authors revealed phenylalanine deaminase (PAD) in the majority of the Citrobacter strains investigated; the activity of PAD varied within a rather considerable range (0.3--4.58 micrometer of phenylpiruvate in 1 hr per 1 mg of bacterial protein). The presence of this enzyme thus served as an auxiliary biochemical test characterising this group of conditionally pathogenic microbes of the Enterobacteriacea family. Tyrosine decarboxylase was absent in 26 of 50 strains of Citrobacter examined. In the rest of the strains the activity of this enzyme was low. Consequently, tyrosine decarboxylase could not be used for identification of microorganisms of the Citrobacter genus.     

Metabolism of dicarboxylic amino acids and their amides in bacteria of the genus Citrobacter]

In 58 Citrobacter strains the pathways of the utilization of dicarbonic amino acids and their amides were studied. These organisms were found to be incapable of decarboxylating glutaminic and asparaginic acids, as well as their amides. All the strains could actively desamidizate asparagine. Not all of these strains showed glutaminase activity. Aspartate-aminotransferase occurred twice as often as alanine-aminotransferase, the level of activity being approximately the same. The Citrobacter strains desamidizated asparaginic acid with great constancy, but only in 1/3 of them this reaction occurred via an aspartase route. The desamidization of asparaginic acid in Citrobacter seemed to proceed in different ways. The desamidization of glutaminic acid was observed only in a part of the strains, and the reaction proceeded less actively.