|Title:||How Does a Ribozyme Work?|
|Group/Series/Folder:||Record Group 8.15 - Institute for Advanced Study|
Series 3 - Audio-visual Materials
|Location:||8.15:3 box 1.5|
|Notes:||Institute for Advanced Study Distinguished Lectures.|
Abstract: The nucleolytic ribozymes catalyse site-specific phosphodiester cleavage and ligation transesterification reactions in RNA. The Varkud satellite (VS) ribozyme is the largest of the nucleolytic ribozymes. In the absence of a crystal structure we have determined the solution structure of the complete ribozyme using small-angle X-ray scattering. The substrate stem-loop docks into the tertiary fold of the ribozyme, where it makes an intimate interaction with the A730 loop, containing the nucleotide A756 which we have found to be important to the function of the ribozyme. A guanine (G638) within the substrate loop of the VS ribozyme also plays a key role in the catalytic activity. Replacement by any other nucleotide result in severe impairment of the cleavage reaction, yet folding of the substrate is not perturbed. Moreover, variant substrates such as G638A bind the ribozyme with similar affinity, acting as competitive inhibitors. We have found that the pH dependence of the ribozyme cleavage reaction is consistent with general acid-base catalysis involving G638 and A756. 5’ phosphorothiolate substitution experiments indicate that G638 is the general base and A756 the general acid for the cleavage reaction. General acid-base catalysis seems to be the basis of all the nucleolytic ribozymes. We show that the intrinsic rate of catalysis by the VS ribozyme is comparable to that of enzymes such as ribonuclease A.
Duration: 74 min.
|Appears in Series:||8.15:3 - Audio-visual Materials|
Videos for Public -- Distinguished Lectures