MODULATION OF THE ACTIVITY OF ANGIOGENIN BY MUTAGENESIS AT ASP-116

Substitution of Asn, Ala or His for Asp-116 in angiogenin increases it s ribonucleolytic activity towards tRNA and, at least in the case of H is, its ability to induce blood-vessel formation (Harper, J.W. and Val lee, B.L. (1988) Proc. Natl. Acad. Sci. USA 85, 7139-7143). Six additi onal Asp-116 mutants have been examined to further probe the basis for this phenomenon. Substitution of Val, Lys, Glu, or Ser increases acti vity towards tRNA 2-, 4-, 9- and 16-fold, respectively, whereas substi tution of Trp and Pro leads to 2- and 10-fold decreases, respectively. Similar changes are seen in activity towards rRNA. Studies of base-cl eavage specificity towards dinucleotide substrates (NpN') reveal a cha nge in preference for G vs. A at the N' position when Ser replaces Asp -116 and a diminished preference for C vs. U at the N position. The Pr o, Lys and Glu mutants have essentially unchanged angiogenic activity. The results demonstrate that the principal effect of replacing Asp-11 6 in angiogenin is to modulate enzymatic activity, possibly through an effect on His-114, and suggest that Asp-116 plays a role in controlli ng specificity.