tailieunhanh - Báo cáo Y học: Arginine 121 is a crucial residue for the specific cytotoxic activity of the ribotoxin a-sarcin

a-Sarcin, a cyclizing ribonuclease secreted by the mould Aspergillus giganteus, is one of the best characterized members of a family of fungal ribotoxins. This protein induces apoptosis in tumour cells due to its highly specific activity on ribosomes. Fungal ribotoxins display a threedimensional protein fold similar to those of a larger group of microbial noncytotoxic RNases, represented by RNases T1 and U2. This similarity involves the three catalytic residues and also the Arg121 residue, whose counterpart in RNase T1, Arg77, is located in the vicinity of the substrate phosphate moiety although its potential functional role is not known. In this. | Eur. J. Biochem. 268 6190-6196 2001 FEBS 2001 Arginine 121 is a crucial residue for the specific cytotoxic activity of the ribotoxin a-sarcin Manuel Masip Javier Lacadena Jose Miguel Mancheno Mercedes Onaderra Antonio Martinez-Ruizt Alvaro Martinez del Pozo and Jose G. Gavilanes Departamento de Bioquimica y Biologia Molecular Facultad de Quimica Universidad Complutense Madrid Spain. a-Sarcin a cyclizing ribonuclease secreted by the mould Aspergillus giganteus is one of the best characterized members of a family of fungal ribotoxins. This protein induces apoptosis in tumour cells due to its highly specific activity on ribosomes. Fungal ribotoxins display a threedimensional protein fold similar to those of a larger group of microbial noncytotoxic RNases represented by RNases T1 and U2. This similarity involves the three catalytic residues and also the Arg121 residue whose counterpart in RNase T1 Arg77 is located in the vicinity of the substrate phosphate moiety although its potential functional role is not known. In this work Arg121 of a-sarcin has been replaced by Gln or Lys. These two mutations do not modify the conformation of the protein but abolish the ribosomeinactivating activity of a-sarcin. In addition the loss of the positive charge at that position produces dramatic changes on the interaction of a-sarcin with phospholipid membranes. It is concluded that Arg121 is a crucial residue for the characteristic cytotoxicity of a-sarcin and presumably of the other fungal ribotoxins. Keywords Aspergillus protein cytotoxin protein-lipid interaction ribotoxin a-sarcin. a-Sarcin is the best characterized member of a family of fungal cytotoxic ribosome-inactivating proteins ribotoxins that cleave one single phosphodiester bond at an evolu-tionarily conserved sequence of the larger rRNA 1 impairing the binding of elongation factors EF-1 and EF-2 2 . This well known exquisite ribonucleolytic action in cell-free systems has been also recently observed in intact cells .