Escherichia coli transforms the leucyl-tRNA synthetase gene in vivo into primordial genes

Abstract

Summary The Last Universal Common Ancestor (LUCA) lived about 4.2 billion years ago and had a nearly modern metabolism1. Genes and proteins must therefore have achieved modern lengths during a time comparable to the error in that date2. How did that happen? We show here thatE colitransformed with a double mutant full-length Leucyl-tRNA synthetase (LeuRS) produces discrete sets of shorter genes. These lack the anticodon-binding domain and large parts of the catalytic domain. They connect remote active site parts in two different ways. Large pre-steady-state bursts confirm that they are the active enzymes. Thesein vivoresults validate earlier designs for ancient aminoacyl-tRNA synthetase enzymes, greatly expanding the sequence space of active synthetases. One construct joins the 56-residue protozyme directly to a 25-residue segment containing the second catalytic signature. It catalyzes both activation and minihelix acylation with amino acids. AlphaFold33predicts that the mRNA sequence encoding the latter fragment is a long hairpin. Thus, 3D structures in the gene itself may promote the deletions4. The deletions appear to reverse the modular evolution of full-length synthetases from simpler catalysts. The reverse evolution we describe could open broad access to primordial gene discovery.