3NKB

A 1.9A crystal structure of the HDV ribozyme precleavage suggests both Lewis acid and general acid mechanisms contribute to phosphodiester cleavage

Summary for 3NKB

• Entry DOI: 10.2210/pdb3nkb/pdb
• Related: 1CX0 1SJ3
• Descriptor: DNA/RNA (5'-D(*(DUR))-D(*GP*G)-R(P*CP*UP*UP*GP*CP*A)-3'), The hepatitis delta virus ribozyme, MAGNESIUM ION, ... (4 entities in total)
• Functional Keywords: catalytic rna, metal-mediated catalysis, phosphodiester cleavage, dna, rna
• Total number of polymer chains: 2
• Total formula weight: 23636.17

Authors

Chen, J.-H.,Yajima, R.,Chadalavada, D.M.,Chase, E.,Bevilacqua, P.C.,Golden, B.L. (deposition date: 2010-06-18, release date: 2010-09-01, Last modification date: 2023-09-06)

Primary citation

Chen, J.H.,Yajima, R.,Chadalavada, D.M.,Chase, E.,Bevilacqua, P.C.,Golden, B.L.
A 1.9 A crystal structure of the HDV ribozyme precleavage suggests both Lewis acid and general acid mechanisms contribute to phosphodiester cleavage.
Biochemistry, 49:6508-6518, 2010

PubMed Abstract: The hepatitis delta virus (HDV) ribozyme and HDV-like ribozymes are self-cleaving RNAs found throughout all kingdoms of life. These RNAs fold into a double-nested pseudoknot structure and cleave RNA, yielding 2',3'-cyclic phosphate and 5'-hydroxyl termini. The active site nucleotide C75 has a pK(a) shifted >2 pH units toward neutrality and has been implicated as a general acid/base in the cleavage reaction. An active site Mg(2+) ion that helps activate the 2'-hydroxyl for nucleophilic attack has been characterized biochemically; however, this ion has not been visualized in any previous structures. To create a snapshot of the ribozyme in a state poised for catalysis, we have crystallized and determined the structure of the HDV ribozyme bound to an inhibitor RNA containing a deoxynucleotide at the cleavage site. This structure includes the wild-type C75 nucleotide and Mg(2+) ions, both of which are required for maximal ribozyme activity. This structure suggests that the position of C75 does not change during the cleavage reaction. A partially hydrated Mg(2+) ion is also found within the active site where it interacts with a newly resolved G.U reverse wobble. Although the inhibitor exhibits crystallographic disorder, we modeled the ribozyme-substrate complex using the conformation of the inhibitor strand observed in the hammerhead ribozyme. This model suggests that the pro-R(P) oxygen of the scissile phosphate and the 2'-hydroxyl nucleophile are inner-sphere ligands to the active site Mg(2+) ion. Thus, the HDV ribozyme may use a combination of metal ion Lewis acid and nucleobase general acid strategies to effect RNA cleavage.

PubMed: 20677830
DOI: 10.1021/bi100670p

Experimental method

X-RAY DIFFRACTION (1.916 Å)