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Single amino acid substitutions affecting the specificity of the fungal ribotoxin mitogillin
Author(s) -
Kao Richard,
Davies Julian
Publication year - 2000
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(99)01753-6
Subject(s) - chemistry , amino acid , biochemistry , biology
Mitogillin and related fungal ribotoxins are small basic ribonucleolytic proteins that inhibit protein synthesis by specifically hydrolyzing a single phosphodiester bond in the universally conserved α‐sarcin/ricin loop (SRL) of large subunit ribosomal RNAs. It was previously shown that mitogillin is a natural derivative of a T1/U2‐like ribonuclease with inserted domains that are involved in target selection and specificity. Site‐directed mutagenesis was used to substitute single amino acids in the previously identified functional domains Ala1–Tyr24 (B1‐L1‐B2 domain) and Lys106–Lys113 (L4 region). Examination of the activities of the mutants in the digestion of polyinosinic acid (a ribonuclease substrate) and specific cleavage of the SRL shows that Asn7Ala and Lys111Gln substitutions lead to altered ribonuclease activity and diminished substrate specificity consistent with the proposed functions of these domains.