Named after Selene, goddess of the moon selenocysteine (Sec) is unique in its unrivaled catalytic power and its insertion into proteins by redefinition of specific UGA stop codons to Sec. In their Communication on page 319 ff., D. Söll, P. O'Donoghue et al. describe the engineering of the Sec-insertion machinery to redefine nearly all 64 codons. Here, the ring-shaped decameric selenocysteine synthase SeIA is essential in providing tRNA-bound Sec as the central building block for selenoprotein translation.

The hyperthermophile Nanoarchaeum equitans is an obligate symbiont growing in coculture with the crenarchaeon Ignicoccus . Ribosomal protein and rRNA-based phylogenies place its branching point early in the archaeal lineage, representing the new archaeal kingdom Nanoarchaeota. The N. equitans genome (490,885 base pairs) encodes the machinery for information processing and repair, but lacks genes for lipid, cofactor, amino acid, or nucleotide biosyntheses. It is the smallest microbial genome sequenced to date, and also one of the most compact, with 95% of the DNA predicted to encode proteins or stable RNAs. Its limited biosynthetic and catabolic capacity indicates that N. equitans ' symbiotic relationship to Ignicoccus is parasitic, making it the only known archaeal parasite. Unlike the small genomes of bacterial parasites that are undergoing reductive evolution, N. equitans has few pseudogenes or extensive regions of noncoding DNA. This organism represents a basal archaeal lineage and has a highly reduced genome.

O-Phosphoserine (Sep), the most abundant phosphoamino acid in the eukaryotic phosphoproteome, is not encoded in the genetic code, but synthesized posttranslationally. Here, we present an engineered system for specific cotranslational Sep incorporation (directed by UAG) into any desired position in a protein by an Escherichia coli strain that harbors a Sep-accepting transfer RNA (tRNA^Sep), its cognate Sep–tRNA synthetase (SepRS), and an engineered EF-Tu (EF-Sep). Expanding the genetic code rested on reengineering EF-Tu to relax its quality-control function and permit Sep-tRNA^Sep binding. To test our system, we synthesized the activated form of human mitogen-activated ERK activating kinase 1 (MEK1) with either one or two Sep residues cotranslationally inserted in their canonical positions (Sep²¹⁸, Sep²²²). This system has general utility in protein engineering, molecular biology, and disease research.

The composition of the human microbiota is recognized as an important factor in human health and disease. Many of our cohabitating microbes belong to phylum-level divisions for which there are no cultivated representatives and are only represented by small subunit rRNA sequences. For one such taxon (SR1), which includes bacteria with elevated abundance in periodontitis, we provide a single-cell genome sequence from a healthy oral sample. SR1 bacteria use a unique genetic code. In-frame TGA (opal) codons are found in most genes (85%), often at loci normally encoding conserved glycine residues. UGA appears not to function as a stop codon and is in equilibrium with the canonical GGN glycine codons, displaying strain-specific variation across the human population. SR1 encodes a divergent tRNA ^Gly _UCA with an opal-decoding anticodon. SR1 glycyl-tRNA synthetase acylates tRNA ^Gly _UCA with glycine in vitro with similar activity compared with normal tRNA ^Gly _UCC . Coexpression of SR1 glycyl-tRNA synthetase and tRNA ^Gly _UCA in Escherichia coli yields significant β-galactosidase activity in vivo from a lacZ gene containing an in-frame TGA codon. Comparative genomic analysis with Human Microbiome Project data revealed that the human body harbors a striking diversity of SR1 bacteria. This is a surprising finding because SR1 is most closely related to bacteria that live in anoxic and thermal environments. Some of these bacteria share common genetic and metabolic features with SR1, including UGA to glycine reassignment and an archaeal-type ribulose-1,5-bisphosphate carboxylase (RubisCO) involved in AMP recycling. UGA codon reassignment renders SR1 genes untranslatable by other bacteria, which impacts horizontal gene transfer within the human microbiota.

The three genes, gatC , gatA , and gatB , which constitute the transcriptional unit of the Bacillus subtilis glutamyl-tRNA ^Gln amidotransferase have been cloned. Expression of this transcriptional unit results in the production of a heterotrimeric protein that has been purified to homogeneity. The enzyme furnishes a means for formation of correctly charged Gln-tRNA ^Gln through the transamidation of misacylated Glu-tRNA ^Gln , functionally replacing the lack of glutaminyl-tRNA synthetase activity in Gram-positive eubacteria, cyanobacteria, Archaea, and organelles. Disruption of this operon is lethal. This demonstrates that transamidation is the only pathway to Gln-tRNA ^Gln in B. subtilis and that glutamyl-tRNA ^Gln amidotransferase is a novel and essential component of the translational apparatus.