endobj 0000116432 00000 n Finally, this finding greatly simplifies and rationalizes the design of fast-cleaving engineered synthetic ribozymes as RNA nucleolytic reagents and as subjects for enzyme kinetics and mechanistic investigations. For this purpose, the two nucleosides around the scissile phosphodiester bond were selectively C-13 labelled in multi-step organic … 0000005948 00000 n 459 0 obj endobj endobj << /N 445 0 R /P 466 0 R /R [ 56 290 288 307 ] /T 437 0 R /V 443 0 R >> endobj 0000096587 00000 n Minimal Hammerhead Ribozymes with Uncompromised Catalytic Activity. 0000044237 00000 n The minimal hammerhead ribozyme is composed of three base paired helices, separated by short linkers of conserved sequence as shown in the crystal structure. << /N 456 0 R /P 49 0 R /R [ 307 53 539 708 ] /T 437 0 R /V 454 0 R >> Molecular Biology, Biology. 0000007632 00000 n 0000004892 00000 n 0000003476 00000 n We use cookies to help provide and enhance our service and tailor content and ads. endobj 0000008237 00000 n Specifically, tertiary interactions form between the loop of stem II and either an internal or terminal loop in … 0000004178 00000 n performs self-cleavage) and therefore cannot catalyze multiple turnovers. endobj endobj 0000003427 00000 n Copyright © 2015 Elsevier Ltd. All rights reserved. 445 0 obj 0000004002 00000 n 0000017833 00000 n 0000004624 00000 n Formation of this additional base-pairing interaction requires only that the substrate RNA sequence contains a U at a position seven nucleotides 3′ to the cleavage site. %%EOF 0000007938 00000 n This small ribozyme is of interest for two reasons. 435 0 obj 0000017298 00000 n << /N 446 0 R /P 466 0 R /R [ 56 290 533 308 ] /T 437 0 R /V 444 0 R >> A minimal hammerhead ribozyme of the present disclosure includes the above structural features of a minimal hammerhead ribozyme, and additionally includes a single tertiary interaction, which tertiary interaction (e.g., a trans-Hoogsteen base pair or similar non-Watson Crick base-pairing) is between a nucleotide in the stem II loop of the catalytic strand and a … The conserved central bases, with few exceptions, are essential for ribozyme’s catalytic activity. https://doi.org/10.1016/j.jmb.2015.05.005. << /Title >> 464 0 obj 0000004534 00000 n 0000018435 00000 n The 2′ … A minimal hammerhead ribozyme construct consists of a conserved core of 15 mostly invariant residues. << /Metadata 385 0 R /Names 386 0 R /OpenAction [ 466 0 R /FitH 1297 ] /Outlines 525 0 R /PageLabels 387 0 R /PageLayout /SinglePage /PageMode /UseOutlines /Pages 384 0 R /StructTreeRoot 388 0 R /Threads [ 437 0 R ] /Type /Catalog >> endobj 446 0 obj flexibility] requirements necessary for hammerhead function, we scrutinized the sequences returned by RNAiFold by measures of deviation (or discrepancy) from structural positional entropy of PLMVd AJ005312.1/282-335. Experiment 3 employs the hammerhead sequence of Experiment 1 with the addition of a 5′-GUU sequence. 463 0 obj 441 0 obj The recent discovery t … 438 0 obj 0000012881 00000 n 0000017236 00000 n 0000003915 00000 n << /Linearized 1 /L 760246 /H [ 4892 426 ] /O 466 /E 117288 /N 8 /T 751426 >> We report here that a single additional trans-Hoogsteen base-pairing interaction in the minimal hammerhead ribozyme transforms an RNA sequence possessing typically modest catalytic activity into one possessing greatly enhanced catalytic activity that is instead typical of full-length natural hammerhead RNAs that have additional extensive tertiary contact interactions. 0000006713 00000 n 0000007020 00000 n It offers a convenient system to study the structure/function relationship of a nucleotide sequence, and is a potential vehicle for the inhibition of gene expression. 458 0 obj Minimal Hammerhead Ribozyme Molecular Biology, Biology. Although the cleavage takes place in the absence of protein enzymes, the hammerhead RNA itself is not a catalyst in its natural state, as it is consumed by the reaction (i.e. 0000100475 00000 n endobj The hammerhead, ribozyme is a small RNA motif that self-cleaves in the presence of a divalent metal ion, generally Mg2+, at a specific phophodiester bond producing 2′,3′-cyclic phosphate and 5′-hydroxyl termini.2,4,6 From: Methods in Enzymology, 2001 The hairpin ribozyme is a small section of RNA that can act as a ribozyme.Like the hammerhead ribozyme it is found in RNA satellites of plant viruses. 0000006406 00000 n The conserved uridine-turn links helix I to helix II and usually contains the sequence CUGA. << /N 450 0 R /P 466 0 R /R [ 301 58 533 275 ] /T 437 0 R /V 446 0 R >> Natural or full-length hammerhead ribozymes are up to 1000-fold more active than their minimal counterparts that lack a complex tertiary interaction that pre-organizes and stabilizes the ribozyme active site, positioning RNA functional groups to facilitate acid-base catalysis. 0000117073 00000 n 0000004802 00000 n 0000096176 00000 n << /N 451 0 R /P 1 0 R /R [ 62 54 295 358 ] /T 437 0 R /V 447 0 R >> One of the prominent features of the full-length hammerhead ribozyme structure (Protein Data Bank (PDB) code, 2GOZ), which is not present in the minimal hammerhead (PDB code, 379D) is that the tertiary stem II loop/stem I bulge interactions appear to induce structural organization of the catalytic core. Helix II and III are linked by a sequence GAAA. << /Filter /FlateDecode /S 201 /O 383 /Length 338 >> 0000099873 00000 n
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