Calcaelin, a new protein with translation-inhibiting, antiproliferative and antimitogenic activities from the mosaic puffball mushroom Calvatia caelata.
Fresh fruiting bodies of the mushroom Calvatia caelata, commonly known as the mosaic puffball, were extracted with 10 mM Tris-HCl buffer (pH 7.2). The extract was subjected to a chromatographic procedure which has been successfully used to isolate ribosome-inactivating proteins from plants. It was first applied to an Affi-gel blue gel column previously equilibrated and eluted with 10 mM Tris-HCl buffer. The adsorbed fraction eluted with 1.5 M NaCl in the buffer was then purified by chromatography on DEAE-cellulose in the same buffer. The unadsorbed fraction yielded essentially a single peak when it was chromatographed on a column of Mono S. When the peak was then gel filtered on Superdex 75, it yielded a homogeneous peak with a molecular mass of 39 kDa. The protein, which was designated calcaelin, was dissociated into two subunits with a molecular mass of 19 kDa and 20 kDa, respectively, in SDS-polyacrylamide gel electrophoresis. The two bands exhibited the same N-terminal amino acid sequence which was somewhat similar to those of plant ribosome-inactivating proteins. A lesser degree of sequence homology to the fungal ribosome inactivating proteins alpha-sarcin and restrictocin was detected. Calcaelin inhibited translation in rabbit reticulocyte lysate with an IC 50 value of 4 nM and displayed a heat-labile RNase activity of 1.58 U/mg toward yeast tRNA. It exhibited an antimitogenic activity toward mouse splenocytes, and it reduced the viability of breast cancer cells. There was no hemagglutinating, antibacterial or antifungal activity.
Ng
Tzi Bun
TB
Department of Biochemistry, Faculty of Medicine, The Chinese University of Hong Kong, Shatin, Hong Kong, PR China. biochemistry@uhk.edu.hk
Lam
Ying Wai
YW
Wang
Hexiang
H
D000818:Animals; D000900:Anti-Bacterial Agents; D000890:Anti-Infective Agents / Q000494:pharmacology; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D001419:Bacteria / Q000187:drug effects; D001487:Basidiomycota; D002455:Cell Division / Q000187:drug effects*; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D008826:Microbial Sensitivity Tests; D008517:Phytotherapy; D010936:Plant Extracts / Q000494:pharmacology*; D010940:Plant Proteins / Q000494:pharmacology*; D011817:Rabbits; D012260:Ribonucleases / Q000378:metabolism; D013154:Spleen / Q000166:cytology / Q000187:drug effects; D014407:Tumor Cells, Cultured / Q000187:drug effects
D000900:Anti-Bacterial Agents; D000890:Anti-Infective Agents; D000972:Antineoplastic Agents, Phytogenic; D010936:Plant Extracts; D010940:Plant Proteins; D012260:Ribonucleases