First simultaneous isolation of a ribosome inactivating protein and an antifungal protein from a mushroom (Lyophyllum shimeji) together with evidence for synergism of their antifungal effects.
From the fruiting bodies of the mushroom Lyophyllum shimeji, a novel ribosome inactivating protein with a molecular weight of 20 kDa and exhibiting antifungal activity against Physalospora piricola (IC(50) = 2.5 microM) and Coprinus comatus was isolated. The protein, designated lyophyllin, was purified by ion exchange chromatography on CM-cellulose, affinity chromatography on Affi-gel Blue Gel, and then ion exchange chromatography on Mono S. Lyophyllin possessed an N-terminal sequence with some similarity to those of plant ribosome-inactivating proteins. It inhibited translation in rabbit reticulocyte lysate with an IC(50) of 1 nM, thymidine uptake by murine splenocytes with an IC(50) of 1 microM and HIV-1 reverse transcriptase activity with an IC(50) of 7.9 nM. Lyophyllin did not manifest ribonuclease or hemagglutinating activity. An antifungal protein, designated Lyophyllum antifungal protein (LAP), with a molecular weight of 14 kDa, and an N-terminal sequence somewhat analogous to those of angiosperm thaumatin-like proteins and thaumatins and an inactive variant of the ubiquitin-conjugating enzyme, was first isolated from Lyophyllum shimeji. LAP was adsorbed on CM-cellulose, Affi-gel blue gel, and Mono S. LAP exerted antifungal activity against P. piricola (IC(50) = 70 nM) and Mycosphaerella arachidicola but not against Rhizoctonia solani, Colletotrichum gossypii, and Coprinus comatus. It exerted very low translation inhibitory activity in a rabbit reticulocyte lysate system (IC(50) = 70 microM) and negligible ribonuclease activity toward yeast transfer RNA and hemagglutinating activity toward rabbit erythrocytes. It inhibited HIV-1 reverse transcriptase with an IC(50) of about 5.2 nM. A synergism in antifungal activities of LAP and lyophyllin against P. piricola was demonstrable.
Lam
S K
SK
Department of Biochemistry, The Chinese University of Hong Kong, Shatin, Hong Kong, China.
Ng
T B
TB
D000363:Agaricales / Q000737:chemistry*; D000595:Amino Acid Sequence; D000935:Antifungal Agents / Q000302:isolation & purification* / Q000494:pharmacology; D003302:Coprinus / Q000187:drug effects; D004357:Drug Synergism; D005656:Fungal Proteins / Q000302:isolation & purification* / Q000494:pharmacology; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D012270:Ribosomes / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid
D000935:Antifungal Agents; D005656:Fungal Proteins; C436962:lyophyllin protein, Lyophyllum shimeji