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The maturase encoded by a group I intron from Aspergillus nidulans stabilizes RNA tertiary structure and promotes rapid splicing.
- Source :
-
Journal of molecular biology [J Mol Biol] 1999 Oct 08; Vol. 292 (5), pp. 987-1001. - Publication Year :
- 1999
-
Abstract
- The AnCOB group I intron from Aspergillus nidulans self-splices, providing the Mg2+ concentration is >/= 15 mM. The splicing reaction is greatly stimulated by a maturase protein encoded within the intron itself. An initial structural and biochemical analysis of the splicing reaction has now been performed. The maturase bound rapidly to the precursor RNA (kon approximately 3 x 10(9) M(-1) min(-1)) and remained tightly bound (koff </= 0.04 min(-1)). The catalytic step of 5' splice-site cleavage occurred at a rate of up to 11 min(-1) under single turnover conditions. The maturase-assisted reaction of heat-denatured RNA proceeded at a rate of about 1 min(-1), arguing that there are early steps of folding that cannot be readily facilitated by the protein. pH analysis revealed a biphasic profile with a pKa of 7.0. The rate of the maturase-assisted reaction was independent of the Mg2+ concentration down to 3 mM. Self-splicing in optimal Mg2+ (>/= 150 mM) was tenfold slower, in part because of the existence of an equilibrium between folded and partially folded RNA. In contrast, the maturase very effectively stabilized tertiary structure in 5 mM Mg2+, a noticeable example being an interaction between the P8 helix and a GNRA sequence that constitutes the L2 terminal loop of the P2 helix. Formation of the 5' splice-site recognition helix was assisted by either the maturase or high concentrations of Mg2+. The maturase was required during splicing so it is not a true chaperone. However, RNase protection assays and kinetic studies suggest that the maturase recognizes and facilitates folding of an intron with limited tertiary structure and even incomplete secondary structure.<br /> (Copyright 1999 Academic Press.)
- Subjects :
- Aspergillus nidulans genetics
Base Sequence
Dose-Response Relationship, Drug
Endopeptidase K metabolism
Guanosine Triphosphate pharmacology
Hot Temperature
Hydrogen-Ion Concentration
Hydrolysis
Kinetics
Magnesium pharmacology
Models, Molecular
Molecular Chaperones physiology
Molecular Sequence Data
Nucleic Acid Conformation drug effects
Nucleic Acid Denaturation
Protein Binding
RNA, Catalytic chemistry
RNA, Catalytic genetics
RNA, Catalytic metabolism
RNA, Fungal genetics
RNA-Directed DNA Polymerase genetics
Ribonuclease T1 metabolism
Aspergillus nidulans enzymology
Introns genetics
RNA Splicing drug effects
RNA, Fungal chemistry
RNA, Fungal metabolism
RNA-Directed DNA Polymerase metabolism
Saccharomyces cerevisiae Proteins
Subjects
Details
- Language :
- English
- ISSN :
- 0022-2836
- Volume :
- 292
- Issue :
- 5
- Database :
- MEDLINE
- Journal :
- Journal of molecular biology
- Publication Type :
- Academic Journal
- Accession number :
- 10512698
- Full Text :
- https://doi.org/10.1006/jmbi.1999.3070