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The relative location of proteins and lipids in particles of medicinal leech salivary gland secretion (SGS) is revealed for the first time. Their sizes and morphology are described. Using scanning electron microscopy and transmission electron microscopy, it was determined that SGS consists of particles of different sizes and form. This picture is supported by confocal laser scanning microscopy of SGS preparations treated with fluorescein isothiocyanate. After incubation with nonionic detergents (Brij 35 and Tween 20), transmission electron microscopy revealed the dissociation of fragments composing protein-lipid particles (PLP), and in this case an increase in free protein concentration determined by a modification of the Lowry method was observed. Perylene probing of lipids in SGS preparations showed that they are concentrated mainly inside PLP and are almost absent on the surface. Cholesterol was detected during SGS probing using the cholesteryl-Bodipy (hydrophobic fluorescent analog of cholesterol) on surface sections during confocal analysis of electron microphotographs of SGS. This analysis detected PLP structures in SGS resembling caveoles full of cholesterol. SGS, preliminary frozen at −70°C, transformed into a multitude of similar small particles visualized by transmission electron microscopy, whose fixed distribution resembled water crystal structure. [ABSTRACT FROM AUTHOR]

The proteins of parasporal inclusions from three Bacillus thuringiensis subspecies (kurstaki, amagiensis, and monterrey) inhibited growth of methanogenic archaea of two species belonging to two genera, Methanobrevibacter arboriphilus and Methanosarcina barkeri. The minimal inhibitory concentrations of these proteins were 20 to 50 μg/ml. Lysozyme exhibited similar bactericidal effect on archaea. The perspective of comparative studies on the effect of polyfunctional proteins on bacteria and archaea is discussed. [ABSTRACT FROM AUTHOR]

Background: Since bactericidal properties of some lysozymes are independent of their glycosidase activity, we have investigated this phenomenon for destabilase-lysozyme (DL) from medicinal leech (Hirudo medicinalis). Methods: Glycosidase activity was determined on Micrococcus luteus, non-enzymatic antibacterial activity of heat-treated DL and of synthetic peptides α1, α2 and α3 (fragments of its primary structure) on M. luteus, Escherichia coli, Bacillus brevis and Streptomyces chrysomallus. Results: Glycosidase activity disappeared after the heating of native DL at 100°C for 40 min. Antibacterial activity of heat-treated DL for M. luteus MDMSU128 and MDMSU140 expressed as minimal inhibitory concentration was 9.8·10–8 and 12·10–8M, respectively, and to E. coli MDMSU52 11·10–8M. Antibacterial activity of synthetic peptide α1 for M. luteus MDMSU128 and for E. coli MDMSU52 was 8.3·10–5 and 4.9·10–5M, respectively. Conclusion: DL is the first invertebrate lysozyme with combined enzymatic and non-enzymatic antibacterial action. Copyright © 2006 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

n expression system for an effective production of recombinant protein Cry9A in bacillary cell has been suggested in the study. The proteins' immunological properties, ability to proteolysis, and biological activity were identical to natural protein. The ability of recombinant Cry9A to form crystal bodies in sporulating cells of Bacillus thuringiensis has been shown. Thus, the first evidences of the fact that Cry-proteins which in natural strains form the crystal bodies together with other endotoxins are able to independently form the crystals has been received. The introduced system including vector replicative carriers, expression cassettes, and a protocol of obtaining and cultivation of strain-producer allows simple manipulations with the gene of delta-endotoxin of Cry9A in gene-engineering experiments. [ABSTRACT FROM AUTHOR]

Preparation and purification of a recombinant protein are described along with characteristics of its specific (for ɛ-(γ-Glu)-Lys and D-dimer substrates) and nonspecific (for L-γ-Glu-pNA) isopeptidase activities; the absence of peptidase function for α-(α-Glu)-Lys substrate is noted. It is shown that the protein exhibits muramidase (cell walls of Micrococcus lysodeikticus) and specific glycosidase activities. The latter was determined towards the fluorogenic substrate 4-methylum-belliferyl-tetra-N-acetyl-β-chitotetraoxide. Antimicrobial activity of recombinant destabilase-lysozyme protein (recDest-Lys) and its 11-membered amphipathic peptide was revealed towards cells of the strict anaerobic Archaean Methanosarcina barkeri, whose cell walls contain no murein. Possible mechanisms of the effect of recDest-Lys on these cells are discussed. [ABSTRACT FROM AUTHOR]