SLIDE 1
O CH2 OH O O P O O O
N1
O CH2 OH O P O O O
N2
O CH2 OH O P O O O
N3
O CH2 OH O P O O O
N4
N A U G C
k =
, , ,
3' - end 5' - end
Na Na Na Na
RNA Structures Stability, Folding and the Role of Hydrogen Bonding - - PowerPoint PPT Presentation
RNA Structures Stability, Folding and the Role of Hydrogen Bonding - - PowerPoint PPT Presentation
RNA Structures Stability, Folding and the Role of Hydrogen Bonding and Protons Peter Schuster Institut fr Theoretische Chemie und Molekulare Strukturbiologie der Universitt Wien Schlo Ringberg, 05.03.2002 5' - end N 1 O The chemical
SLIDE 2
SLIDE 3
Structural Constraints and Hydrogen Bonding in RNA
Single stranded RNA molecules form structures, which combine double-helical stacking (A-type) regions with loops and metal ion (Mg2 ) coordinated centers.
SLIDE 4
The three-dimensional structure of a short double helical stack
SLIDE 5
Canonical Watson-Crick base pairs: cytosine – guanine uracil – adenine
W.Saenger, Principles of Nucleic Acid Structure, Springer, Berlin 1984
SLIDE 6
O O O O O H H H H H H H H H H H N N N N N N N N N N N O O H N N H O N N N N N N N
G=U G C
- U=G
Canonical Watson-Crick base-pair Wobble base-pairs
Wobble base pairs in RNA double-helical stacks
SLIDE 7
C G ``A´´ U
2,6-diamino purine 2-keto, 6-amino purine 2,6-diketo purine 5-keto, 7-amino, 1,6,8-triaza indolicine 5- , 7- , 1,6,8-triaza indolicine amino keto 2-amino,6-keto purine 2-keto, 4-amino pyrimidine
2- , 4- pyrimidine amino keto
2,4-di pyrimidine keto 2,6-diamin pyrimidine
- 2-
, 6-keto pyrazine amino 2- , 6- pyrazine keto amino
Color code: Donor—Acceptor Acceptor—Donor
Hydrogen bonding patterns for Watson- Crick base pairs
S.A. Benner et al., Reading the palimpsest: Contemporary biochemical data and the RNA world. In: R.F.Gesteland and J.F.Atkins, eds. The RNA World, pp.27-70. CSHL Press, 1993
SLIDE 8
Classification of purine- pyrimidine base pairs
SLIDE 9
Classification of purine-purine base pairs
SLIDE 10
Classification of pyrimidine- pyrimidine base pairs
SLIDE 11
General classification
- f base pairs
N.B.Leontis and E. Westhof, RNA 7:499-512 (2001)
SLIDE 12
Stacking of heterocyclic aromatic molecules without sugar-phosphate backbone
Example: N6,N9-dimethyl adenine, D. Pörschke and F. Eggers, Eur.J.Biochem. 26:490-498 (1972)
SLIDE 13
Stacking of RNA single strands
Example: poly-A, D.Pörschke. Elementary steps of base recognition and helix-coil transitions in nucleic acids. In: I.Pecht and R.Rigler, eds. Chemical Relaxation in Molecular Biology, pp.191-218. Springer-Verlag, Berlin 1977.
SLIDE 14
Three-dimensional structure of phenylalanyl-transfer-RNA
SLIDE 15
RNA Secondary Structures and their Properties
RNA secondary structures are listings of Watson-Crick and GU wobble base pairs, which are free of knots and pseudokots. Secondary structures are folding intermediates in the formation of full three-dimensional structures.
D.Thirumalai, N.Lee, S.A.Woodson, and D.K.Klimov. Annu.Rev.Phys.Chem. 52:751-762 (2001)
SLIDE 16
5'-End 5'-End 5'-End 3'-End 3'-End 3'-End
70 60 50 40 30 20 10 GCGGAU AUUCGC UUA AGDDGGGA M CUGAAYA AGMUC TPCGAUC A ACCA GCUC GAGC CCAGA UCUGG CUGUG CACAG
Sequence Secondary Structure Symbolic Notation
Definition of the secondary structure of phenylalanyl-tRNA
SLIDE 17
5.10
2
2.90
8 14 15 18
2.60
17 23 19 27 22 38 45 25 36 33 39 40
3.10
43
3.40
41
3.30 7.40
5 3 7
3.00
4 10 9
3.40
6 13 12
3.10
11 21 20 16 28 29 26 30 32 42 46 44 24 35 34 37 49
2.80
31 47 48
S0 S1
Kinetic Structures Free Energy S0 S0 S1 S2 S3 S4 S5 S6 S7 S8 S10 S9 Minimum Free Energy Structure Suboptimal Structures T = 0 K , t T > 0 K , t T > 0 K , t finite
5.90
Different notions of RNA structure
SLIDE 18
RNA Minimum Free Energy Structures
Efficient algorithms based on dynamical programming are available for computation of secondary structures for given
- sequences. Inverse folding algorithms compute sequences
for given secondary structures.
M.Zuker and P.Stiegler. Nucleic Acids Res. 9:133-148 (1981) Vienna RNA Package: http:www.tbi.univie.ac.at (includes inverse folding, suboptimal structures, kinetic folding, etc.) I.L.Hofacker, W. Fontana, P.F.Stadler, L.S.Bonhoeffer, M.Tacker, and P. Schuster. Mh.Chem. 125:167-188 (1994)
SLIDE 19
UUUAGCCAGCGCGAGUCGUGCGGACGGGGUUAUCUCUGUCGGGCUAGGGCGC GUGAGCGCGGGGCACAGUUUCUCAAGGAUGUAAGUUUUUGCCGUUUAUCUGG UUAGCGAGAGAGGAGGCUUCUAGACCCAGCUCUCUGGGUCGUUGCUGAUGCG CAUUGGUGCUAAUGAUAUUAGGGCUGUAUUCCUGUAUAGCGAUCAGUGUCCG GUAGGCCCUCUUGACAUAAGAUUUUUCCAAUGGUGGGAGAUGGCCAUUGCAG
Criterion of Minimum Free Energy
Sequence Space Shape Space
SLIDE 20
Sk I. = ( ) ψ
- Gk
k
= ( ) f S
Sequence space Shape space Non-negative numbers
Mapping from sequence space into phenotype space and into free energies
SLIDE 21
.... GC UC .... CA .... GC UC .... GU .... GC UC .... GA .... GC UC .... CU
d =1
H
d =1
H
d =2
H
Point mutations as moves in sequence space
SLIDE 22
λj = 27 ,
/
12 λk = (k)
j
| | Gk
λ κ
cr = 1 - -1 (
1)
/ κ- λ λ
k cr . . . .
> λ λ
k cr . . . .
< Network is connected Gk Network is connected not Gk Connectivity Threshold: Alphabet Size : = 4
- AUGC
G S S
k k k
= ( ) | ( ) =
- 1
- I
I
j j
- cr
2 0.5 3 0.4226 4 0.3700
Mean degree of neutrality and connectivity of neutral networks
SLIDE 23
A connected neutral network
SLIDE 24
Giant Component
A multi-component neutral network
SLIDE 25
Kinetic Folding of RNA at Elementary Step Resolution
The RNA folding process is resolved to base pair closure, base pair cleavage and base pair shift. The kinetic folding behavior is determined by computation
- f a sufficiently large ensemble of individual folding trajectories and taking an
average over them. The folding behavior is illustrated by barrier trees showing the path of lowest energy between two local minima of free energy.
C.Flamm, W.Fontana, I.L.Hofacker and P.Schuster. RNA, 6:325-338 (2000)
SLIDE 26
closure shift cleavage
Move set for elementary steps in kinetic RNA folding
SLIDE 27
Folding dynamics of the sequence GGCCCCUUUGGGGGCCAGACCCCUAAAAAGGGUC
SLIDE 28
C U G G G A A A A A U C C C C A G A C C G G G G G U U U C C C C G G G G G G G G G G G G G G G G G G G G C C C C C C C C U U U U U U G G G G G C C C C C C C C C C C C C U U U A A A A A A A A A A U
3’-end
Minimum free energy conformation S0 Suboptimal conformation S1
C G
One sequence is compatible with two structures
SLIDE 29
Sh S1
(h)
S6
(h)
S7
(h)
S5
(h)
S2
(h)
S9
(h)
Free energy G Local minimum Suboptimal conformations
Search for local minima in conformation space
SLIDE 30
Free energy G0
- Free energy G0
- "Reaction coordinate"
Sk Sk S S Saddle point T
- k
T
- k
"Barrier tree"
SLIDE 31
5.10
2
2.90
8 14 15 18
2.60
17 23 19 27 22 38 45 25 36 33 39 40
3.10
43
3.40
41
3.30 7.40
5 3 7
3.00
4 10 9
3.40
6 13 12
3.10
11 21 20 16 28 29 26 30 32 42 46 44 24 35 34 37 49
2.80
31 47 48
S0 S1
Barrier tree of a sequence with two conformations
5.90
SLIDE 32
A ribozyme switch
E.A.Schultes, D.B.Bartel, One sequence, two ribozymes: Implication for the emergence of new ribozyme folds. Science 289 (2000), 448-452
SLIDE 33
U U U U U G G G G G G G G G G G G G G G G G A A A A A A A A A A C C C C C C C C C C C C C C C
Cleavage site
The "hammerhead" ribozyme
OH OH OH ppp 5' 5' 3' 3'
The smallest known catalytically active RNA molecule
SLIDE 34
Two ribozymes of chain lengths n = 88 nucleotides: An artificial ligase (A) and a natural cleavage ribozyme of hepatitis-
- virus (B)
SLIDE 35
The sequence at the intersection: An RNA molecules which is 88 nucleotides long and can form both structures
SLIDE 36
Reference for the definition of the intersection and the proof of the intersection theorem
SLIDE 37
Two neutral walks through sequence space with conservation of structure and catalytic activity
SLIDE 38
Sequence of mutants from the intersection to both reference ribozymes
SLIDE 39
Reference for postulation and in silico verification of neutral networks
SLIDE 40