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Protein-DNA Computation by Stochastic Assembly Cascade
Roy Bar-Ziv, Tsvi Tlusty, and Albert Libchaber
PNAS Sep.3, 2002
Presented by Je-Gun Joung
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Overview
• The assembly of RecA on single-stranded DAN is measured and interpreted as a stochastic finite-state machine
• This machine is able to discriminate fine differences between sequences
• In vivo stochastic biomolecular computation based on low-specificity protein-DNA binding
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Molecular Machine by Stochastic Assembly Cascade
• Hardware– A test tube filled with a solution of single-stranded DN
A molecule, RecA protein, and ATP molecules
– Output device (molecular machine reporters): a change in the rotational motion of the DNA molecule
• Software– Two basic operation: nucleation and disassembly
– Chang the machine’s internal state (the current length of RecA)
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Molecular Machine by Stochastic Assembly Cascade
RecA
ATP
ssDNA
Hardware
Output device(fluorescence anisotropy)
QN
QN-1
Q2
Q1
Q0
……
Two basic operation (nucleation and disassembly)
Software
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RecA• The protein responsible for recombination in bacteria
5’ssDNA
RecA coats ssDNA
Alignment with homologous DNA sequence
Formation of joint chromosomal DNA
3’
5’5’3’
3’
RecA
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RecA
RecA/Rad51 DNA Filaments
RecA Monomer
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Biochemistry of RecA Assembly
TAC TAC TAC TAC TAC TAC TAC TAC TAC TAC TAC TAC TAC
5’ 3’
• Nucleation (assembly): extend the filament by long chunks
TAC TAC TAC TAC TAC TAC TAC TAC TAC TAC TAC TAC TAC5’ 3’
• Disassembly: removes monomers one by one
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Simulation of a 26-state Stochastic Assembly Cascade
QN
QN-1
Q2
Q1
Q0
……
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A Deterministic Reading Head Changes its Internal State
x e8 x d8 d7 x x d6 d5 x e9 d9 x
Q6
QN QN-1 Q2 Q1 Q0………
………
dn : disassembly from state Qn
en : nucleation to state Qn
x : noting happens
Tape
Reading Head
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A Simple Set of Rules
Suppose that the machine is at state Qn
• If it read dn, it moves to state Qn+1
• If it read em and m < n the machine moves to state Qm
• In all other cased, if it read x and dm with m n, or em with m n, then it stays at stateoves to state Qn
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“Sequence-detector” Machine
• Assembly cascades on single-stranded DNAs on 39 or 78 bases long (13- and 26-stage machines)
• Infer its internal dynamics from observable output• A sufficient set of observations to overcome the n
oisiness of the output 105-106 fluctuating DNA-RacA complexes• A very smooth ensemble-average signal
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An Exponential Sensitivity of the Assembly Cascade
• Both curves decay exponentially close to saturation with a higher slope for the longer sequence that has more cascade stages
• A: fluorescence anisotryopy
• S: the normalized anisotropy
• Open circles: (TAC)13
• Triangles: (TAC)26
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Sequence-Discrimination Capability
Very similar triplet repeats Localized differences in nonuniform sequences
the difference between the fluorescence anisotropy signal
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Discussion
• By choice of other types of sequence base vectors, the stochastic cascade machinery can encode and decode mixtures in terms of other transforms
• Additional operations to manipulate sequences such as recombination
• The question of whether RecA assembly is used for natural computation requires in vivo testing.