A CORE FOLDING MODEL FOR CATALYSIS BY THE HAMMERHEAD RIBOZYME ACCOUNTS FOR ITS EXTRAORDINARY SENSITIVITY TO ABASIC MUTATIONS

Introducing abasic nucleotides at each of 13 positions in the conserve d core of the hammerhead ribozyme causes a large decrease in the exten t of catalysis [Peracchi, A., et al. (1996) Proc. Natl. Acad. Sci. U.S .A. 93, 11522]. This extreme sensitivity to structural defects is in c ontrast to the behavior of protein enzymes and larger ribozymes. Sever al additional differences in the behavior of the hammerhead relative t o that of protein enzymes and larger ribozymes are described herein. T he deleterious effects of the abasic mutations are not relieved by low ering the temperature, by increasing the concentration of monovalent o r divalent metal ions, or by adding polyamines, in contrast to effects observed with protein enzymes and large RNA enzymes. In addition, the abasic mutations do not significantly weaken substrate binding. These results and previous observations are all; accounted for by a ''core folding'' model in which the stable ground state structure of the hamm erhead ribozyme complexed with the substrate is a partially folded sta te that must undergo an additional folding event to achieve its cataly tic conformation. We propose that the peculiar behavior of the hammerh ead arises because the limited structural interconnections in a small RNA enzyme do not allow the ground state to stably adopt the catalytic conformation; within the globally folded catalytic conformation, limi ted structural interconnections may further impair catalysis by hamper ing the precise alignment of active site functional groups. This behav ior represents a basic manifestation of the well-recognized interconne ction between folding and catalysis.