Linked Life Data

15169770

Source:http://linkedlifedata.com/resource/pubmed/id/15169770

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
31
pubmed:dateCreated
2004-7-26
pubmed:abstractText
Engineered RNase P ribozymes are promising gene-targeting agents that can be used in both basic research and clinical applications. We have previously selected ribozyme variants for their activity in cleaving an mRNA substrate from a pool of ribozymes containing randomized sequences. In this study, one of the variants was used to target the mRNA encoding thymidine kinase (TK) of herpes simplex virus 1 (HSV-1). The variant exhibited enhanced cleavage and substrate binding and was at least 30 times more efficient in cleaving TK mRNA in vitro than the ribozyme derived from the wild type sequence. Our results provide the first direct evidence to suggest that a point mutation at nucleotide 95 of RNase P catalytic RNA from Escherichia coli (G(95) --> U(95)) increases the rate of cleavage, whereas another mutation at nucleotide 200 (A(200) --> C(200)) enhances substrate binding of the ribozyme. A reduction of about 99% in TK expression was observed in cells expressing the variant, whereas a 70% reduction was found in cells expressing the ribozyme derived from the wild type sequence. Thus, the RNase P ribozyme variant is highly effective in inhibiting HSV-1 gene expression. Our study demonstrates that ribozyme variants increase their cleavage activity and efficacy in blocking gene expression in cells through enhanced substrate binding and rate of cleavage. These results also provide insights into the mechanism of how RNase P ribozymes efficiently cleave an mRNA substrate and, furthermore, facilitate the development of highly active RNase P ribozymes for gene-targeting applications.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jul
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
30
pubmed:volume
279
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
32063-70
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed-meshheading:15169770-Base Sequence, pubmed-meshheading:15169770-Blotting, Northern, pubmed-meshheading:15169770-Blotting, Western, pubmed-meshheading:15169770-Catalysis, pubmed-meshheading:15169770-Escherichia coli, pubmed-meshheading:15169770-Humans, pubmed-meshheading:15169770-Kinetics, pubmed-meshheading:15169770-Molecular Sequence Data, pubmed-meshheading:15169770-Nucleic Acid Conformation, pubmed-meshheading:15169770-Plasmids, pubmed-meshheading:15169770-Point Mutation, pubmed-meshheading:15169770-Protein Binding, pubmed-meshheading:15169770-RNA, pubmed-meshheading:15169770-RNA, Catalytic, pubmed-meshheading:15169770-RNA, Messenger, pubmed-meshheading:15169770-Retroviridae, pubmed-meshheading:15169770-Ribonuclease P, pubmed-meshheading:15169770-Simplexvirus, pubmed-meshheading:15169770-Thymidine Kinase, pubmed-meshheading:15169770-Time Factors, pubmed-meshheading:15169770-Viral Proteins
pubmed:year
2004
pubmed:articleTitle
Engineered RNase P ribozymes increase their cleavage activities and efficacies in inhibiting viral gene expression in cells by enhancing the rate of cleavage and binding of the target mRNA.
pubmed:affiliation
Program in Infectious Diseases and Immunity, Program in Comparative, Biochemistry, School of Public Health, University of California, Berkeley, 94720, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S., Research Support, Non-U.S. Gov't