United States of America, As Represented by the Secretary of Agriculture

Beatriz Martinez Fernandez

David M Donovan

Lorena Rodriguez Rubio

Pilar Garcia Suarez

Ana Rodriguez

Virion-associated peptidoglycan hydrolases have a potential as antimicrobial agents due to their ability to lyse Gram positive bacteria on contact. Fusion proteins HydH5SH3b and HydH5Lyso comprising full-length peptidoglycan hydrolase HydH5 from thebacteriophage vB_SauS-phi-IPLA88 in combination with the SH3b cell wall-binding domain from lysostaphin or full length lysostaphin, respectively, exhibited high lytic activity against livecells. CHAPSH3b, a HydH5 CHAP (cysteine, histidine-dependent amidohydrolase/peptidase) domain from truncated HydH5 in combination with the SH3b domain of lysostaphin, exhibited the highest lytic activity against livecells. HydH5 and its derivative fusions lysed bovine and human, methicillin-resistant(MRSA) N315 strain, and humanstrains in zymogram, plate lysis and turbidity reduction assays. HydH5 and its derivative fusions displayed antimicrobial synergy with the endolysin LysH5 (also encoded by vB_SauS-phi-IPLA88) in vitro suggesting the two enzymes have distinct cut sites and thus may be more efficient in combination for elimination of staphylococcal infections.

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Virion-associated peptidoglycan hydrolases have a potential as antimicrobial agents due to their ability to lyse Gram positive bacteria on contact. Full-length HydH5, a virion-associated peptidoglycan hydrolase from thebacteriophage vB_SauS-phi-IPLA88, and two truncated derivatives, containing only the CHAP domain, exhibited high lytic activity against livecells. Three different fusion proteins were created and showed higher staphylolytic activity than the parental enzyme or its deletion construct. Parental and fusion proteins lysedcells in zymograms, plate lysis and turbidity reduction assays. In plate lysis assays, HydH5 and its derivative fusions lysed bovine and humanMRSA N315 strain, and humanstrains. HydH5 and its derivative fusions proteins displayed antimicrobial synergy with the endolysin LysH5 in vitro suggesting that the two enzymes have distinct cut sites and thus may be more efficient in combination for the elimination of staphylococcal infections.

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Austrias, Spain

Beatriz Martinez Fernandez