title: Mutagenesis of the novel Hericium erinaceus ribonuclease, RNase He1, reveals critical responsible residues for enzyme stability and activity.
abstract: Here, we determined the sequence of a cDNA encoding a guanylic acid-specific ribonuclease (RNase He1) from Hericium erinaceus that exhibits high sequence identity (59%) with RNase Po1, an enzyme with anti-cancer activity and which is found in Pleurotus ostreatus. RNase He1 and RNase Po1 have similar structures and heat stabilities; hence, RNase He1 may also have potential as an anti-cancer agent. Therefore, we initiated structure-function studies to further characterize the enzyme. Based on the RNase Po1 structure, RNase He1 is predicted to form 3 disulfide bonds involving Cys7-Cys98, Cys5-Cys83, and Cys47-Cys81 linkages. The Cys5Ala mutant exhibited no RNase activity, whereas the Cys81Ala mutant retained RNase activity, but had reduced heat stability. Therefore, the Cys5-Cys83 bond in RNase He1 is essential for the structure of the RNase active site region. Similarly, the Cys47-Cys81 bond helps maintain the conformational stability of the active site region, and may contribute to the greater heat stability of RNase He1.
authors.affiliation: School of Pharmacy, Nihon University.
authors.forename: Hiroko
authors.initials: H
authors.lastname: Kobayashi
authors.forename: Naomi
authors.initials: N
authors.lastname: Motoyoshi
authors.forename: Tadashi
authors.initials: T
authors.lastname: Itagaki
authors.forename: Norio
authors.initials: N
authors.lastname: Inokuchi
mesh_terms: D001487:Basidiomycota / Q000201:enzymology*; D049109:Cell Proliferation / Q000187:drug effects; D003545:Cysteine / Q000235:genetics; D004795:Enzyme Stability; D018922:HL-60 Cells; D006801:Humans; D016296:Mutagenesis; D012313:RNA / Q000378:metabolism; D012260:Ribonucleases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology
chemical_list: D012313:RNA; D012260:Ribonucleases; D003545:Cysteine