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Kinetic and secondary structure analysis of Naegleria andersoni GIR1, a group I ribozyme whose putative biological function is site-specific hydrolysis.
Biochemistry. 1997 Dec 23; 36(51):16345-54.B

Abstract

NanGIR1 is a catalytic element inserted in the P6 loop of a group I intron (NanGIR2) in the small subunit rRNA precursor of the protist Naegleria andersoni [Einvik, C., Decatur, W. A., Embley, T. M., Vogt, V. M., and Johansen, S. (1997) RNA 3, 710-720]. It catalyzes site-specific hydrolysis at an internal processing site (IPS) after a G residue that immediately follows the P9 stem-loop. Functional and structural analyses were initiated to compare NanGIR1 to group I introns that carry out self-splicing. Chemical modification and site-directed mutagenesis studies showed that NanGIR1 shares many structural elements with other group I introns, but also contains a pseudoknot (P15), which is important for catalytic activity. Deletion analysis revealed the boundaries of the minimum self-cleaving unit (178 nucleotides). The rate of self-cleavage was measured as a function of mono- and divalent ion concentration, temperature, and pH. The reaction at the IPS yields 5'-phosphate and 3'-hydroxyl termini, requires Mg2+or Mn2+ ions, and is first-order in [OH-] between pH 5.0 and 8.5. The latter results suggest that the nucleophile in the reaction is hydroxide or possibly a Mg2+-coordinated hydroxide. With a second-order rate constant of 1 x 10(5) min-1 M-1, the self-cleavage reaction of NanGIR1 is 2 orders of magnitude faster than a similar site-specific hydrolysis reaction of the circular form of the Tetrahymena group I intron.

Authors+Show Affiliations

Department of Chemistry and Biochemistry, Howard Hughes Medical Institute, University of Colorado, Boulder, Colorado 80309-0215, USA.No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

9405070

Citation

Jabri, E, et al. "Kinetic and Secondary Structure Analysis of Naegleria Andersoni GIR1, a Group I Ribozyme Whose Putative Biological Function Is Site-specific Hydrolysis." Biochemistry, vol. 36, no. 51, 1997, pp. 16345-54.
Jabri E, Aigner S, Cech TR. Kinetic and secondary structure analysis of Naegleria andersoni GIR1, a group I ribozyme whose putative biological function is site-specific hydrolysis. Biochemistry. 1997;36(51):16345-54.
Jabri, E., Aigner, S., & Cech, T. R. (1997). Kinetic and secondary structure analysis of Naegleria andersoni GIR1, a group I ribozyme whose putative biological function is site-specific hydrolysis. Biochemistry, 36(51), 16345-54.
Jabri E, Aigner S, Cech TR. Kinetic and Secondary Structure Analysis of Naegleria Andersoni GIR1, a Group I Ribozyme Whose Putative Biological Function Is Site-specific Hydrolysis. Biochemistry. 1997 Dec 23;36(51):16345-54. PubMed PMID: 9405070.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Kinetic and secondary structure analysis of Naegleria andersoni GIR1, a group I ribozyme whose putative biological function is site-specific hydrolysis. AU - Jabri,E, AU - Aigner,S, AU - Cech,T R, PY - 1998/1/24/pubmed PY - 1998/1/24/medline PY - 1998/1/24/entrez SP - 16345 EP - 54 JF - Biochemistry JO - Biochemistry VL - 36 IS - 51 N2 - NanGIR1 is a catalytic element inserted in the P6 loop of a group I intron (NanGIR2) in the small subunit rRNA precursor of the protist Naegleria andersoni [Einvik, C., Decatur, W. A., Embley, T. M., Vogt, V. M., and Johansen, S. (1997) RNA 3, 710-720]. It catalyzes site-specific hydrolysis at an internal processing site (IPS) after a G residue that immediately follows the P9 stem-loop. Functional and structural analyses were initiated to compare NanGIR1 to group I introns that carry out self-splicing. Chemical modification and site-directed mutagenesis studies showed that NanGIR1 shares many structural elements with other group I introns, but also contains a pseudoknot (P15), which is important for catalytic activity. Deletion analysis revealed the boundaries of the minimum self-cleaving unit (178 nucleotides). The rate of self-cleavage was measured as a function of mono- and divalent ion concentration, temperature, and pH. The reaction at the IPS yields 5'-phosphate and 3'-hydroxyl termini, requires Mg2+or Mn2+ ions, and is first-order in [OH-] between pH 5.0 and 8.5. The latter results suggest that the nucleophile in the reaction is hydroxide or possibly a Mg2+-coordinated hydroxide. With a second-order rate constant of 1 x 10(5) min-1 M-1, the self-cleavage reaction of NanGIR1 is 2 orders of magnitude faster than a similar site-specific hydrolysis reaction of the circular form of the Tetrahymena group I intron. SN - 0006-2960 UR - https://www.unboundmedicine.com/medline/citation/9405070/Kinetic_and_secondary_structure_analysis_of_Naegleria_andersoni_GIR1_a_group_I_ribozyme_whose_putative_biological_function_is_site_specific_hydrolysis_ L2 - https://doi.org/10.1021/bi9718595 DB - PRIME DP - Unbound Medicine ER -