title: Minimized natural versions of fungal ribotoxins show improved active site plasticity.
abstract: Fungal ribotoxins are highly specific extracellular RNases which cleave a single phosphodiester bond at the ribosomal sarcin-ricin loop, inhibiting protein biosynthesis by interfering with elongation factors. Most ribotoxins show high degree of conservation, with similar sizes and amino acid sequence identities above 85%. Only two exceptions are known: hirsutellin A and anisoplin, produced by the entomopathogenic fungi Hirsutella thompsonii and Metarhizium anisopliae, respectively. Both proteins are similar but smaller than the other known ribotoxins (130 vs 150 amino acids), displaying only about 25% sequence identity with them. They can be considered minimized natural versions of their larger counterparts, best represented by α-sarcin. The conserved α-sarcin active site residue Tyr48 has been replaced by the geometrically equivalent Asp, present in the minimized ribotoxins, to produce and characterize the corresponding mutant. As a control, the inverse anisoplin mutant (D43Y) has been also studied. The results show how the smaller versions of ribotoxins represent an optimum compromise among conformational freedom, stability, specificity, and active-site plasticity which allow these toxic proteins to accommodate the characteristic abilities of ribotoxins into a shorter amino acid sequence and more stable structure of intermediate size between that of other nontoxic fungal RNases and previously known larger ribotoxins.
authors.affiliation: Departamento de Bioquímica y Biología Molecular I, Facultad de Ciencias Químicas, Universidad Complutense, 28040 Madrid, Spain.
authors.forename: Moisés
authors.initials: M
authors.lastname: Maestro-López
authors.affiliation: Departamento de Bioquímica y Biología Molecular I, Facultad de Ciencias Químicas, Universidad Complutense, 28040 Madrid, Spain.
authors.forename: Miriam
authors.initials: M
authors.lastname: Olombrada
authors.affiliation: Departamento de Bioquímica y Biología Molecular I, Facultad de Ciencias Químicas, Universidad Complutense, 28040 Madrid, Spain.
authors.forename: Lucía
authors.initials: L
authors.lastname: García-Ortega
authors.affiliation: Departamento de Bioquímica y Biología Molecular I, Facultad de Ciencias Químicas, Universidad Complutense, 28040 Madrid, Spain.
authors.forename: Daniel
authors.initials: D
authors.lastname: Serrano-González
authors.affiliation: Departamento de Bioquímica y Biología Molecular I, Facultad de Ciencias Químicas, Universidad Complutense, 28040 Madrid, Spain.
authors.forename: Javier
authors.initials: J
authors.lastname: Lacadena
authors.affiliation: Departamento de Bioquímica y Biología Molecular I, Facultad de Ciencias Químicas, Universidad Complutense, 28040 Madrid, Spain.
authors.forename: Mercedes
authors.initials: M
authors.lastname: Oñaderra
authors.affiliation: Departamento de Bioquímica y Biología Molecular I, Facultad de Ciencias Químicas, Universidad Complutense, 28040 Madrid, Spain.
authors.forename: José G
authors.initials: JG
authors.lastname: Gavilanes
authors.affiliation: Departamento de Bioquímica y Biología Molecular I, Facultad de Ciencias Químicas, Universidad Complutense, 28040 Madrid, Spain. Electronic address: alvaromp@quim.ucm.es.
authors.forename: Álvaro
authors.initials: Á
authors.lastname: Martínez-Del-Pozo
mesh_terms: D020134:Catalytic Domain; D004722:Endoribonucleases / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry*; D005658:Fungi / Q000201:enzymology*; D052981:Metarhizium / Q000201:enzymology*; D009154:Mutation; D010445:Peptide Elongation Factors / Q000737:chemistry; D014176:Protein Biosynthesis; D011487:Protein Conformation; D012260:Ribonucleases / Q000737:chemistry*; D012270:Ribosomes / Q000378:metabolism; D013056:Spectrophotometry, Ultraviolet; D014443:Tyrosine / Q000737:chemistry
chemical_list: D005656:Fungal Proteins; D010445:Peptide Elongation Factors; C014801:alpha-sarcin; D014443:Tyrosine; D004722:Endoribonucleases; D012260:Ribonucleases
keywords: Anisoplin; Hirsutellin; Insecticidal; RNases; Sarcin