ENGINEERING PROTEIN-BASED MACHINES TO EMULATE KEY STEPS OF METABOLISM(BIOLOGICAL ENERGY-CONVERSION)

Metabolism is the conversion of available energy sources to those ener gy forms required for sustaining and propagating living organisms; thi s is simply biological energy conversion. Proteins are the machines of metabolism; they are the engines of motility and the other machines t hat interconvert energy forms not involving motion. Accordingly, metab olic engineering becomes the use of natural protein-based machines for the good of society. In addition, metabolic engineering can utilize t he principles, whereby proteins function, to design new protein-based machines to fulfill roles for society that proteins have never been ca lled upon throughout evolution to fulfill. This article presents argum ents for a universal mechanism whereby proteins perform their diverse energy conversions; it begins with background information, and then as serts a set of five axioms for protein folding, assembly, and function and for protein engineering. The key process is the hydrophobic foldi ng and assembly transition exhibited by properly balanced amphiphilic protein sequences. The fundamental molecular process is the competitio n for hydration between hydrophobic and polar, e.g., charged, residues . This competition determines T-t, the onset temperature for the hydro phobic folding and assembly transition, N-hh, the numbers of waters of hydrophobic hydration, and the pKa of ionizable functions. Reported a cid-base titrations and pH dependence of microwave dielectric relaxati on data simultaneously demonstrate the interdependence of T-t, N-hh an d the pKa using a series of microbially prepared protein-based poly(30 mers) with one glutamic acid residue per 30mer and with an increasing number of more hydrophobic phenylalanine residues replacing valine res idues. Also, reduction of nicotinamides and flavins is shown to lower T-t, i.e., to increase hydrophobicity.Furthermore, the argument is pre sented, and related to an extended Henderson-Hasselbalch equation, whe rein reduction of nicotinamides represents an increase in hydrophobici ty and resulting hydrophobic-induced pKa shifts become the basis for u nderstanding a primary energy conversion (proton transport) process of mitochondria. (C) 1998 John Wiley & Sons, Inc.