title: Ribonuclease T1 is active when both catalytic histidines are replaced by aspartate.
abstract: The enzymatic activity of many ribonucleases (RNases) depends on two histidines, as in RNase A, or one histidine and/or glutamate, as in microbial RNases belonging to the T1 family. In RNase T1, substitution of either one or both of the two histidines at positions 40 and 92 by a variety of other amino acids reduces the activity more than 100-fold. However, the double variant His40-->Asp/His92-->Asp retains a significant residual enzymatic activity towards RNA and guanylyl-3',5'-cytidine, indicating that a pair of substituted side chains in these positions, both with acid functionality, can confer enzymatic activity. It was shown that the substitution of histidine with glutamate in the variant His40-->Glu yields an enzyme with drastically reduced activity and leads to inactivation in the His92-->Glu, His40-->Glu/His92-->Glu variants. For the variants where histidine is substituted with aspartate we found measurable activity from 1% (His40-->Asp) to 6% (His40-->Glu/His92-->Asp) towards RNA.
authors.affiliation: Institut für Kristallographie, Freie Universität Berlin, Germany.
authors.forename: O
authors.initials: O
authors.lastname: Landt
authors.forename: J
authors.initials: J
authors.lastname: Thölke
authors.forename: H P
authors.initials: HP
authors.lastname: Grunert
authors.forename: W
authors.initials: W
authors.lastname: Saenger
authors.forename: U
authors.initials: U
authors.lastname: Hahn
mesh_terms: D001224:Aspartic Acid / Q000737:chemistry* / Q000235:genetics; D001665:Binding Sites / Q000235:genetics; D002384:Catalysis; D004789:Enzyme Activation / Q000235:genetics; D006639:Histidine / Q000737:chemistry* / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D006163:Ribonuclease T1 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D013329:Structure-Activity Relationship
chemical_list: D001224:Aspartic Acid; D006639:Histidine; D006163:Ribonuclease T1