[Single-chain, low molecular weight kininogen in the blood plasma of rabbits: isolation and fragmentation by porcine pancreatic kallikrein]. / Odnotsepochechnyi nizkomolekuliarnyi kininogen plazmy krovi krolika: vydelenie i kharakter fragmentatsii pankreaticheskim kallikreinom svin'i.

A highly purified preparation of low molecular weight kininogen (LMrK) was isolated from the plasminogen-free rabbit blood plasma, using chromatography on DEAE-Sepharose CL-6B, gel filtration on Ultrogel AcA 34 and Sephadex G-100 as well as gradient chromatography on a hydroxylapatite column. The yield of the 320-fold purified LMrK was 16%. Trypsin released 13-14 micrograms-eq. of bradykinin (BK) from 1 mg of LMrK or 0.85-0,95 mol of BK per mol of kininogen. Rabbit LMrK consists of one polypeptide chain of Mr 69 000 and pI 4.63. Porcine pancreatic kallikrein splits off kinin from the LMrK polypeptide chain by disrupting two peptide bonds resulting in the formation of S-S-bound two chain molecule. After reduction of the S-S bonds by dithioerithritol the latter is separated into a heavy (Mr 61 000) and light (Mr 6 800) chains. A biologically active peptide was isolated from the products of CNBr cleavage of LMrK. This peptide consists of Lys-BK elongated from the C-terminal with several amino acid residues. Rabbit LMrK closely resembles human LMrK in terms of Mr, pI and location of the kinin fragment in the protein molecule.

[Enzymes incorporated in polyelectrolyte complexes. Effect of matrix conformational changes and phase transitions in solutions on catalytic properties]. / Fermenty, vkliuchennye v poliélektrolitnye kompleksy. Vliianie konformatsionnykh izmenenii matritsy i fazovykh perekhodov v rastvorakh na kataliticheskie svoistva.

Immobilization of enzymes (penicillin amidase and alpha-chymotrypsin) in water-soluble nonstoichiometric polyeloctrolyte complexes (PEC) formed by poly(4-vinyl-N-ethylpyridinium bromide) (polycation) and polymethacrylic acid (polyanion) was carried out. Particles of these PEC consist of a nucleus formed by sequences of salt bonds between the units of oppositely charged polyelectrolytes and the hydrophylic shell formed by ionized groups of polyanions which is in excess in PEC. Such a structure of PEC particles results in a cooperative phase transitions of these systems at slight variations of pH and ionic strength. The work demonstrates phase diagrams of PEC solutions. The values of pH and ionic strength at which phase transitions in solutions of different PEC occur were elucidated. The decrease of pH value from 6.1 to 5.7 leads to reversible phase transition followed by a saltatory increase of Km for immobilized penicillin amidase by 5-10 fold depending on substrate used. The phase transition induced by ionic strength increase up to 0,27 M NaCl doesn't change significantly the Km-value of enzymic reaction. The phenomenon observed can be accounted for by the different structure of PEC particles. The catalytic properties of immobilized chymotrypsin were shown to depend on the loci of enzyme attachment. If the enzyme is bound to polyanion, neither conformational changes of the matrix nor phase transition in solution influence its accessibility for the protein inhibitor, but rather change the binding constant. If the enzyme is attached to polycation, i.e. included in the polycomplex nucleus, two fractions of enzymes accessible and inaccessible for protein inhibitor appear.(ABSTRACT TRUNCATED AT 250 WORDS)