Antibacterial Non-Glycosidase Activity of Invertebrate Destabilase-Lysozyme and of Its Helical Amphipathic Peptides

Abstract

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^–8 M, 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.

Lysozymes in the animal kingdom

“…Xue et al (2007) reported two i-type lysozymes in the eastern oyster, both of which signifi cantly inhibited the growth of E. coli, Vibrio vulnifi cus and Pediococcus cerevisiae, but at different concentrations. Zavalova et al (2006) detected high antimicrobial activity of heattreated destabilase/lysozyme that lacked glycosidase activity towards both Micrococcus luteus and E. coli. They considered this as evidence for a non-enzymatic antibacterial mode of action of the destabilase/lysozyme from medicinal leech besides its enzymatic activity.…”

cDNA cloning and tissue expression of plasma lysozyme in the eastern oyster, Crassostrea virginica

“…Antifungal and antiprotozoal activities of lysozymes have also been reported and have been attributed to the hydrolysis of N-acetylglucosamine linkages in chitin that makes up the cell wall of fungus and some protozoa [3e5]. There is also increasing evidence that lysozymes kill Gram-positive and Gram-negative bacteria by nonenzymatic means [6,7]. While the antimicrobial activity of lysozymes has repeatedly been demonstrated in vitro, the contribution of lysozymes to host defences is only beginning to be unravelled in vivo [8,9].…”

Highly active fractions of the medicinal leech recombinant destabilase-lysozyme

“…Фермент Дест-Лиз (КФ 3.5.1.44), компонент секрета слюнных клеток медицинской пиявки, сначала был обнаружен и выделен как эндо-e-(g-Glu)-Lys-изопептидаза, расщепляющая эти связи в стабилизированном фибрине -основе кровяного тромба и в продукте его протеолиза -D-димере [1,2]. Позднее у этого фермента были выявлены лизоцимная и прямая антимикробная функции [3,4]. В результате экспрессии одного из трёх генов Дест-Лиз был получен и очищен его рекомбинантный вариант, которому свойственны все функции природной формы [5].…”