Ssutton: PTL Logic

Introduction

I am working to develop a new type of logic, called Post-Translational Logic, or PTL. PTL devices regulate the post-translational modifications of proteins to define system state and control cell function.

Current synthetic biological circuits make use of protein-DNA and RNA-RNA interactions to control gene expression in bacteria-- such circuits are examples of Transcrition-based Logic. A brief comparison of the two types of logic is as follows:

Transcription-based Logic

• Engineered around gene expression
• Typical parts: transcriptional regulators, translational regulators
• Typical signal: PoPS, resulting in desired cellular concentrations of proteins.
• Easier to engineer than PTL
• Slow response time (hours)
• Uses one subset of cellular functions

PTL

• Engineered around protein modifications
• Typical parts: kinases, phosphorylation sites, docking sites
• Typical signal: rate of modification, resulting in desired state of proteins.
• More difficult to engineer than Transcription-based logic.
• Fast response time (seconds)
• Explores a new set of applications

In designing PTL logic, I am working to answer the following questions:

• What is a PTL part?
• What is a PTL device?
• What signals are passed between devices?
• What are device performance specifications?

Below I will describe some of my ideas.

A Basic PTL System: Kinase-Kinase circuit

The most intuitive definition of a PTL device is illustrated as follows.

• In this system, each device is essentially a MAP Kinase. Each device shown here contains the following parts:
  • Kinase Scaffold
  • Docking Site
  • PO₄ Site
  • Docking Groove
  • PO₄ Groove
  • Catalytic Domain
• The above parts are needed so that one MAP Kinase can recognize another. MAP Kinases recognize their substrates in two ways:
  • The Docking Groove of the upstream kinase (e.g. Device1) binds to the Docking Site of the downstream kinase (e.g. Device2).
  • The PO₄ Groove of the upstream kinase binds to the PO₄ Site of the downstream kinase.
• Once a MAP Kinase PO₄ Site has been phosphorylated, the MAP Kinase undergoes a conformational change which positions the Catalytic Domain into its active conformation with catalytic activity parameter k_cat.
• Devices can be linearly arranges so as to form a cascade:
  • Device1 phosphorylates and activates Device2.
  • Device2 phosphorylates and activates Device3, etc.
• The signal passed between devices is some sort of catalytic activity of one kinase for another.

The problem with the above system is that the output of one device can only be received by a device that contains compatible Docking and PO4 parts. The figure below illustrates points of part-part interactions, with red line connecting parts that must be compatibile.

This trans-part compatibility defeats the purpose of a universal signal carrier, and limits the utility and versatility of PTL devices. Note that we encountered the same problem when defining PDL devices and signals.

A solution is to re-draw the device boundaries such that all corresponding parts are within the same devices:

Device 1 could be added upstream of any other device, and device part would be phosphorylated with maximal rate Vmax1.

Other types of devices: Localization

Phosphorylation controls many intracellular behaviors, including:

• allosteric activation (described by the MAP Kinases above) or inhibition
• translocation
• degradation
• protein complex formation
• other binding events

One of the largest challenges facing PTL is finding a signal carrier that can descrbie all of the above functions.

As an example, let's examine nuclear localization:

Thus far, I have shown two useful signal carriers: Vmax and Vnuc. Let's examine how the two signal carriers could be integrated into one circuit.

Combining Vmax and Vnuc into one circuit

Here is an example of a kinase that modulates the rate of transport of a Protein into the nucleus:

We could imagine a circuit in which an input device phosphorylates a downstream device on two Phosphorylation sites, thus resulting in two distinctive output signals:

Other types signals we might want to define and measure