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Bio-Design Automation EE5393 – University of Minnesota Brian’s Automated Modular Biochemical Instantiator ECE What, Expense How, Why, Reports … What are we doing? • Investigating design strategies for generating “netlists” of protein-protein biochemical reactions. How are we going about it? • Applying circuit CAD methodologies: modularity / abstractions / hierarchical designs. Why are we bothering? • Such tools and methods will revolutionize the way synthetic biology is done. Playing by the Rules Playing by the Rules Rules for integrated circuits: amplifier v1 1 0 rin1 1 0 9e12 rjump 1 4 1e-12 rin2 4 0 9e12 e1 3 0 1 2 999k e2 6 0 4 5 999k e3 9 0 8 7 999k rload 9 0 10k r1 2 3 10k rgain 2 5 10k r2 5 6 10k r3 3 7 10k r4 7 9 10k r5 6 8 10k r6 8 0 10k .dc v1 0 10 1 .print dc v(9) .end circuit netlist SPICE waveforms Playing by the Rules Rules for biochemistry: Gillespie’s SPICE SSA X=100, Y = 30 Xa = Xb = Xn= 0 Y =0 biochemical reactions and initial quantities of proteins histogram: resulting quantities of proteins Playing by the Rules Rules for biochemistry: Gillespie’s SPICE SSA algorithms widely studied data structures (Gibson & Bruck, Fett & Riedel); approximation methods (Petzold); hybrid discrete/continuous methods (Kaznessis); … dynamics well studied mathematics (Tyson, Khammash, Doyle, …); biology (Arkin, Endy, Brent); … computation (Winfree, Shapiro); … Gillespie’s SPICE SSA X=100, Y = 30 Xa = Xb = Xn= 0 Y =0 Biochemical Netlists Netlists found in nature: • Elucidated by biologists. X=100, Y = 30 Xa = Xb = Xn= 0 Y =0 New Netlists: • Designed by skilled experimentalists (by tinkering with existing mechanisms). Where does the netlist come from? Synthetic Biology • Positioned as an engineering discipline. – “Novel functionality through design”. – Repositories of standardized parts. • Driven by experimental expertise in particular domains of biology. – Modify gene regulation, signaling pathways, metabolic pathways… Building Bridges "Think of how engineers build bridges. They design quantitative models to help them understand what sorts of pressure and weight the bridge can withstand, and then use these equations to improve the actual physical model. [In our work on memory in yeast cells] we really did the same thing.” – Pam Silver, Harvard 2007 Engineering Design • Quantitative modeling. • Mathematical analysis. • Incremental and iterative design changes. Synthetic Biology Feats of synthetic bio-engineering: • Cellulosic ethanol (Nancy Ho, Purdue, ’04) • Anti-malarial drugs (Jay Keasling, UC Berkeley, ‘06) • Tumor detection (Chris Voigt, UCSF ‘06) Strategy: apply experimental expertise; formulate ad-hoc designs; perform extensive simulations. Building Digital Circuits inputs outputs x1 f1 ( x1 ,K, xm) f 2 ( x1 ,K, xm) x2 .. . xm digital circuit .. . f n ( x1 ,K, xm ) • Design is driven by the input/output specification. • CAD tools are not part of the design process; they are the design process. [computational] [computational] Synthetic Analysis Biology “There are known ‘knowns’; and there are unknown ‘unknowns’; but today I’ll speak of the known ‘unknowns’.” – Donald Rumsfeld, 2004 Molecular Inputs Known / Known Unknown Biological Process Molecular Products Given Unknown Unknown Known Gene Regulation Hard to tinker with this; but it’s the way computation is done… Biochemistry in a Nutshell Nucleotides: { A, C , T , G} DNA: string of n nucleotides (n ≈ 109) ... ACCGTTGAATGACG... Amino acid: coded by a sequence of 3 nucleotides. { A, C , T , G }3 {a1 ,K , a20 } Proteins: produced from a sequence of m amino acids (m ≈ 103) called a “gene”. {a1 ,K , a20 } m protein Custom Gene Synthesis Going from reading genetic codes to writing them. US Patent 20070122826 (pending): “The present invention relates to a minimal set of protein-coding genes which provides the information required for replication of a free-living organism in a rich bacterial culture medium.” – J. Craig Venter Institute Custom Gene Synthesis Going from reading genetic codes to writing them. Moderator: “Some people have accused you of playing God.” J. Craig Venter: “Oh no, we’re not playing.” Biochemical Netlists What are we doing? • Figuring out how to design netlists in terms of abstract protein types so that we meet desired specs. X=100, Y = 30 Xa = Xb = Xn= 0 Y =0 Why? • Implement computation & signal processing on protein quantities. Ok, but how? Playing by the Rules Biochemical Reactions: how types of molecules combine. 2a + + b c Biochemical Reactions + species count cell 9 8 6 5 7 9 Discrete chemical kinetics; spatial homogeneity. Biochemical Reactions Relative rates or (reaction propensities): slow + medium + fast + Discrete chemical kinetics; spatial homogeneity. Stochastic Kinetics The probability that a given reaction is the next to fire is proportional to: • Its rate. • The number of ways that the reactants can combine. R1 R2 R3 k1 2A B 3 C k2 B 2C k3 3A A C 2B See D. Gillespie, “Stochastic Chemical Kinetics”, 2006. Stochastic Kinetics For each reaction Ri let k n1X1 n2 X2 X1 X2 i k n1 n2 Choose the next reaction according to: i Pr( Ri ) j j Design Automation for Integrated Circuits Behavioral Specification (e.g., DSP function) Register Level Design Structural Description (e.g., memory and functional units) Logic Synthesis Circuit-Level Description (e.g., NAND2 and D flip-flops) SPICE waveforms Design Automation for Integrated Biochemistry Circuits Behavioral Specification (e.g., DSP function) Register Level Design Structural Description (e.g., memory and functional units) Logic Biochemical Synthesis Biochemical Netlist (e.g., Proteins, Enzymes) SPICE STA Engine waveforms PSB 2009: “Stochastic Transient Analysis Biochemical Systems” Design Automation for Integrated Biochemistry Circuits Behavioral Specification (e.g., DSP function) Register Level Design Structural Description (e.g., memory and functional units) Logic Biochemical Synthesis Biochemical Netlist (e.g., Proteins, Enzymes) SPICE STA Engine DAC 07, SB 3.0: “The Synthesis of Stochastic Biochemical Systems” waveforms Design Automation for Integrated Biochemistry Circuits Behavioral Specification (e.g., DSP function) Register Level Design Joint work with Keshab Parhi’s group. Structural Description (e.g., memory and functional units) Biochemical Synthesis Biochemical Netlist (e.g., Proteins, Enzymes) SPICE STA Engine Brian’s Automated Modular Biochemical Instantiator waveforms