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Journal of Biomolecular Structure and Dynamics Publication details, including instructions for authors and subscription information: http://www.tandfonline.com/loi/tbsd20
Secondary Structures of Tetrahymena Thermophila rRNA IVS Sequence Involved in Its Self-Splicing Reactions: A New Computer Analysis a
a
Giorgio Benedetti , Pasquale De Santis & Stefano Morosetti
a
a
Department of Chemistry , University of Rome I , P.le A. Moro 5, 00185 , Rome , Italy Published online: 21 May 2012.
To cite this article: Giorgio Benedetti , Pasquale De Santis & Stefano Morosetti (1990) Secondary Structures of Tetrahymena Thermophila rRNA IVS Sequence Involved in Its Self-Splicing Reactions: A New Computer Analysis, Journal of Biomolecular Structure and Dynamics, 7:6, 1269-1277, DOI: 10.1080/07391102.1990.10508564 To link to this article: http://dx.doi.org/10.1080/07391102.1990.10508564
PLEASE SCROLL DOWN FOR ARTICLE Taylor & Francis makes every effort to ensure the accuracy of all the information (the “Content”) contained in the publications on our platform. However, Taylor & Francis, our agents, and our licensors make no representations or warranties whatsoever as to the accuracy, completeness, or suitability for any purpose of the Content. Any opinions and views expressed in this publication are the opinions and views of the authors, and are not the views of or endorsed by Taylor & Francis. The accuracy of the Content should not be relied upon and should be independently verified with primary sources of information. Taylor and Francis shall not be liable for any losses, actions, claims, proceedings, demands, costs, expenses, damages, and other liabilities whatsoever or howsoever caused arising directly or
indirectly in connection with, in relation to or arising out of the use of the Content.
Downloaded by [New York University] at 01:37 08 February 2015
This article may be used for research, teaching, and private study purposes. Any substantial or systematic reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any form to anyone is expressly forbidden. Terms & Conditions of access and use can be found at http://www.tandfonline.com/page/terms-and-conditions
Journal ofBiomolecular Structure & Dynamics, ISSN 0739-1102 Volume 7, Issue Number 6 (1990), «~Adenine Press (1990).
Secondary Structures of Tetrahymena Thermophila rRNA IVS Sequence Involved in Its Self-Splicing Reactions: ANew Computer Analysis
Downloaded by [New York University] at 01:37 08 February 2015
Giorgio Benedetti, Pasquale DeSantis and Stefano Morosetti Department of Chemistry University of Rome I P.leA Moro 5 00185 Rome, Italy Abstract The secondary structures of Tetrahymena thermophila rRNA IVS sequence involved in the self-splicing reactions, are theoretically investigated with a refined computer method previously proposed, able to select a set of the deepest free energy RNA secondary structures under constraints of model hypotheses and experimental evidences. The secondary structures obtained are characterized by the close proximity of self-reactions sites and account for double mutations experiments, and differential digestion data.
Introduction The importance of Tetrahymena thermophila rRNA IVS molecule resides mainly in its self-splicing ability, and enzymatic activity (1,2). The secondary structure has been appealed to explain its various autocatalytic reactions (3-5). Yet some problems are present; in particular it is not clear how the 3' end ofiVS is held in position to react with the cyclization site following U 15 (3). The existence of structurally discriminating experimental data makes computer algorithms, able to take such evidences into account, more suitable to search of secondary structures. We had already developed a computer method ofthis kind (6), here furtherly expanded: we also introduced model hypotheses in the algorithm; we used the new approach so to obtain IVS secondary structures with sites of self-reactions in close proximity. The main features of the computer algorithm were: 1) the ability of selecting a set of optimal free energy secondary structures with computer time requirements proportional to N 2 ; 2) the possibility of introducing an adjustable arrangement between the experimental data respect and the free energy content of the structure. This graduality is useful because the most stable secondary structure might not correspond to the biological meaningful one as a consequence of tertiary and environmental effects
1269
Benedetti eta/.
1270
(7); moreover other secondary structures should be invoked in order to completely describe the biological phenomenon.
Methods
Downloaded by [New York University] at 01:37 08 February 2015
The Introduction of Model Hypotheses It is often possible to express hypotheses which relate the biological activity of RNA sequence to the secondary structure ofits specific domains. See the following examples: a) in the case ofTrp operon (8) a fragment of the sequence is thought to remain as a single strand so to interact with a protein. Therefore it has not to be available for the formation of a double strand secondary structure; b) on the contrary, a double strand is present in the hypothesis of control in the primer formation ofCoiEl plasmid replication (9).
We have thus developed a method able to introduce structural proposals expressed as single and/or double strand evidences, following the approach used in our previous paper (6) for the experimental data. We put the term experimental data in addition to the total free energy, as: N Nb l(il:" 1 slot= ce {s(m) + 2 L su+k(i)J} m=l i=l j=O
:L
:L
where S101 is the experimental contribution, N is the number of nucleotides in the sequence, Nb is the total number of the helical regions in the structure, l(i) and k(i) are respectively the length and the starting position of the helical region i. Ce is the reference value corresponding to the mean free energy stacking of the considered sequence. s depends on the experimental evidences, and it is attributed to any base by adding the numbers which represent the enzymatic cleavage at the contiguous phosphorous, provided the absolute maximum value Is Imust be 1. We assigned + 1 for single strand and -1 for double strand evidence. A difference between major and minor enzymatic cleavage activity is often reported, and in this case we attribute ± 1/2 to the minor sites. If a larger spectrum of experimental values is present, the whole 0 + 1 interval can be utilized. Here we define an additional term, ~01 , to the total free energy; it depends likewise S101 on the model hypotheses considered. The final expression of~01, obtained like slot• is:
~ot=1
Nb l(il:- 1
N
L
m=l
r(m) + 2
L L
i=l
r[j+k(i)]
j=O
where r(m) = + 1 if the m h base is single strand and r(m) = -1 if it is double. If the m 1h base is not involved in the model hypotheses considered, the r(m) value is 0.
1271
Secondary Structures of Tetrahymena rRNA Finally, the global function E101 attributed to any structure, is:
where: G 101 is the free energy contribution, A, is the coefficient used to weight the experimental term in comparison with the energetic one, and~ is the coefficient for the model term.
Downloaded by [New York University] at 01:37 08 February 2015
The terms of E101 can be grouped in a more suitable way. Indeed G 101 can be explicited as:
or: Nb Gtot =
L
Gs(i) +
m=l where Gs
Journal of Biomolecular Structure and Dynamics Publication details, including instructions for authors and subscription information: http://www.tandfonline.com/loi/tbsd20
Secondary Structures of Tetrahymena Thermophila rRNA IVS Sequence Involved in Its Self-Splicing Reactions: A New Computer Analysis a
a
Giorgio Benedetti , Pasquale De Santis & Stefano Morosetti
a
a
Department of Chemistry , University of Rome I , P.le A. Moro 5, 00185 , Rome , Italy Published online: 21 May 2012.
To cite this article: Giorgio Benedetti , Pasquale De Santis & Stefano Morosetti (1990) Secondary Structures of Tetrahymena Thermophila rRNA IVS Sequence Involved in Its Self-Splicing Reactions: A New Computer Analysis, Journal of Biomolecular Structure and Dynamics, 7:6, 1269-1277, DOI: 10.1080/07391102.1990.10508564 To link to this article: http://dx.doi.org/10.1080/07391102.1990.10508564
PLEASE SCROLL DOWN FOR ARTICLE Taylor & Francis makes every effort to ensure the accuracy of all the information (the “Content”) contained in the publications on our platform. However, Taylor & Francis, our agents, and our licensors make no representations or warranties whatsoever as to the accuracy, completeness, or suitability for any purpose of the Content. Any opinions and views expressed in this publication are the opinions and views of the authors, and are not the views of or endorsed by Taylor & Francis. The accuracy of the Content should not be relied upon and should be independently verified with primary sources of information. Taylor and Francis shall not be liable for any losses, actions, claims, proceedings, demands, costs, expenses, damages, and other liabilities whatsoever or howsoever caused arising directly or
indirectly in connection with, in relation to or arising out of the use of the Content.
Downloaded by [New York University] at 01:37 08 February 2015
This article may be used for research, teaching, and private study purposes. Any substantial or systematic reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any form to anyone is expressly forbidden. Terms & Conditions of access and use can be found at http://www.tandfonline.com/page/terms-and-conditions
Journal ofBiomolecular Structure & Dynamics, ISSN 0739-1102 Volume 7, Issue Number 6 (1990), «~Adenine Press (1990).
Secondary Structures of Tetrahymena Thermophila rRNA IVS Sequence Involved in Its Self-Splicing Reactions: ANew Computer Analysis
Downloaded by [New York University] at 01:37 08 February 2015
Giorgio Benedetti, Pasquale DeSantis and Stefano Morosetti Department of Chemistry University of Rome I P.leA Moro 5 00185 Rome, Italy Abstract The secondary structures of Tetrahymena thermophila rRNA IVS sequence involved in the self-splicing reactions, are theoretically investigated with a refined computer method previously proposed, able to select a set of the deepest free energy RNA secondary structures under constraints of model hypotheses and experimental evidences. The secondary structures obtained are characterized by the close proximity of self-reactions sites and account for double mutations experiments, and differential digestion data.
Introduction The importance of Tetrahymena thermophila rRNA IVS molecule resides mainly in its self-splicing ability, and enzymatic activity (1,2). The secondary structure has been appealed to explain its various autocatalytic reactions (3-5). Yet some problems are present; in particular it is not clear how the 3' end ofiVS is held in position to react with the cyclization site following U 15 (3). The existence of structurally discriminating experimental data makes computer algorithms, able to take such evidences into account, more suitable to search of secondary structures. We had already developed a computer method ofthis kind (6), here furtherly expanded: we also introduced model hypotheses in the algorithm; we used the new approach so to obtain IVS secondary structures with sites of self-reactions in close proximity. The main features of the computer algorithm were: 1) the ability of selecting a set of optimal free energy secondary structures with computer time requirements proportional to N 2 ; 2) the possibility of introducing an adjustable arrangement between the experimental data respect and the free energy content of the structure. This graduality is useful because the most stable secondary structure might not correspond to the biological meaningful one as a consequence of tertiary and environmental effects
1269
Benedetti eta/.
1270
(7); moreover other secondary structures should be invoked in order to completely describe the biological phenomenon.
Methods
Downloaded by [New York University] at 01:37 08 February 2015
The Introduction of Model Hypotheses It is often possible to express hypotheses which relate the biological activity of RNA sequence to the secondary structure ofits specific domains. See the following examples: a) in the case ofTrp operon (8) a fragment of the sequence is thought to remain as a single strand so to interact with a protein. Therefore it has not to be available for the formation of a double strand secondary structure; b) on the contrary, a double strand is present in the hypothesis of control in the primer formation ofCoiEl plasmid replication (9).
We have thus developed a method able to introduce structural proposals expressed as single and/or double strand evidences, following the approach used in our previous paper (6) for the experimental data. We put the term experimental data in addition to the total free energy, as: N Nb l(il:" 1 slot= ce {s(m) + 2 L su+k(i)J} m=l i=l j=O
:L
:L
where S101 is the experimental contribution, N is the number of nucleotides in the sequence, Nb is the total number of the helical regions in the structure, l(i) and k(i) are respectively the length and the starting position of the helical region i. Ce is the reference value corresponding to the mean free energy stacking of the considered sequence. s depends on the experimental evidences, and it is attributed to any base by adding the numbers which represent the enzymatic cleavage at the contiguous phosphorous, provided the absolute maximum value Is Imust be 1. We assigned + 1 for single strand and -1 for double strand evidence. A difference between major and minor enzymatic cleavage activity is often reported, and in this case we attribute ± 1/2 to the minor sites. If a larger spectrum of experimental values is present, the whole 0 + 1 interval can be utilized. Here we define an additional term, ~01 , to the total free energy; it depends likewise S101 on the model hypotheses considered. The final expression of~01, obtained like slot• is:
~ot=1
Nb l(il:- 1
N
L
m=l
r(m) + 2
L L
i=l
r[j+k(i)]
j=O
where r(m) = + 1 if the m h base is single strand and r(m) = -1 if it is double. If the m 1h base is not involved in the model hypotheses considered, the r(m) value is 0.
1271
Secondary Structures of Tetrahymena rRNA Finally, the global function E101 attributed to any structure, is:
where: G 101 is the free energy contribution, A, is the coefficient used to weight the experimental term in comparison with the energetic one, and~ is the coefficient for the model term.
Downloaded by [New York University] at 01:37 08 February 2015
The terms of E101 can be grouped in a more suitable way. Indeed G 101 can be explicited as:
or: Nb Gtot =
L
Gs(i) +
m=l where Gs
