Engineering synthetic signaling proteins with ultrasensitive input/output control

Abstract

Many signaling proteins are built from simple, modular components, yet display highly complex signal-processing behavior. Here we explore how modular domains can be used to build an ultrasensitive switch--a nonlinear input/output function that is central to many complex biological behaviors. By systematically altering the number and affinity of modular autoinhibitory interactions, we show that we can predictably convert a simple linear signaling protein into an ultrasensitive switch.

Metabolic engineering of microorganisms for biofuels production: from bugs to synthetic biology to fuels

“…Another thrust of synthetic biology is the creation of reusable parts that have useful and predictable behavior. The development of a diverse array of regulatable expression systems [ 54,55] that can be used to fine-tune expression is very useful in engineering metabolic pathways. Also, switches that sense and respond to environmental changes [ 56,57] and can subsequently initiate down-stream regulation have exciting opportunities for biofuels production.…”

Proligands with protease‐regulated binding activity identified from cell‐displayed prodomain libraries

“…1,2 Proteins and peptides can undergo conformation and activity changes in response to changes in pH or temperature, small-molecule binding, 3,4 or enzymatic activity. 5 Such sophisticated input/output control has proven useful in the development of biosensors, 6,7 cell internalizing agents, 5 and selective in vivo diagnostics and therapeutics. 8 More specifically, the localization and biological activity of proproteins including prohormones and proenzymes is frequently regulated by site-specific proteolytic cleavage leading to conformational rearrangement 9 or active-site exposure.…”

Using the Population‐Shift Mechanism to Rationally Introduce “Hill‐type” Cooperativity into a Normally Non‐Cooperative Receptor

“…[21][22][23] Cooperativity, also called homotropic allostery (all sites bind identical molecules), thus differs from heterotropic allostery (sites bind different molecules), which instead alters the placement (i.e., midpoint) of the binding curve without changing its underlying shape. While extensive literature exists regarding the design of heterotropically allosteric receptors, [24][25][26] the rational design of cooperativity has seen relatively little success, [11][12][13][14][15][16] perhaps because details of the mechanism render its design rather non-intuitive. First, the all-or-nothing effect of cooperativity requires the creation of systems in which a higher affinity site is occupied only after a lower affinity site that binds the same ligand is already filled.…”

“…[7,8] Allosteric cooperativity, however, which is arguably the simplest solution to this problem, [9,10] has seen adaptation to only a handful of small-molecule [11,12] and biopolymer-based receptors. [13][14][15][16] Here we explore and articulate design principles underlying this mechanism by engineering it into a normally noncooperative receptor, thus improving the receptor s ability to respond to subtle concentration changes.The occupancy of an allosterically cooperative receptor goes aswhere K Half is the concentration at which half of all binding sites are occupied and n H , the "Hill coefficient," provides a convenient metric of cooperativity: a system is noncooperative at n H = 1, and approaches maximum cooperativity as n H approaches the number of binding sites on the receptor. [17] (Note: here we discuss positive cooperativity, which steepens the binding curve.…”

“…[7,8] Allosteric cooperativity, however, which is arguably the simplest solution to this problem, [9,10] has seen adaptation to only a handful of small-molecule [11,12] and biopolymer-based receptors. [13][14][15][16] Here we explore and articulate design principles underlying this mechanism by engineering it into a normally noncooperative receptor, thus improving the receptor s ability to respond to subtle concentration changes.…”