SH3 and OB are the simplest, oldest and most common protein domains within the translation system. SH3 and OB domains are β-barrels that are structurally similar but are topologically distinct. To transform an OB domain to a SH3 domain, β-strands must be permuted in a multistep and evolutionarily implausible mechanism. Here, we explored relationships between SH3 and OB domains of ribosomal proteins, initiation and elongation factors using a combined sequence- and structure-based approach. We detect a common core of SH3 and OB domains, as a region of significant structure and sequence similarity. The common core contains four β-strands and a loop, but omits the fifth β-strand, which is variable and is absent from some OB and SH3 domain proteins. The structure of the common core immediately suggests a simple permutation mechanism for interconversion between SH3 and OB domains, which appear to share an ancestor. The OB domain was formed by duplication and adaptation of the SH3 domain, or vice versa, in a simple and probable transformation. By employing the folding algorithm AlphaFold2, we demonstrated that an ancestral reconstruction of a permuted SH3 sequence folds into an OB structure, and an ancestral reconstruction of a permuted OB sequence folds into a SH3 structure. The tandem SH3 and OB domains in the universal ribosomal protein uL2 share a common ancestor, suggesting that the divergence of these two domains occurred before the Last Universal Common Ancestor.

Hemerythrin-like proteins have generally been studied for their ability to reversibly bind oxygen through their binuclear nonheme iron centers. However, in recent years, it has become increasingly evident that some members of the hemerythrin-like superfamily also participate in many other biological processes. For instance, the binuclear nonheme iron site of YtfE, a hemerythrin-like protein involved in the repair of iron centers in Escherichia coli, catalyzes the reduction of nitric oxide to nitrous oxide, and the human F-box/LRR-repeat protein 5, which contains a hemerythrin-like domain, is involved in intracellular iron homeostasis. Furthermore, structural data on hemerythrin-like domains from two proteins of unknown function, PF0695 from Pyrococcus furiosus and NMB1532 from Neisseria meningitidis, show that the cation-binding sites, typical of hemerythrin, can be absent or be occupied by metal ions other than iron. To systematically investigate this functional and structural diversity of the hemerythrin-like superfamily, we have collected hemerythrin-like sequences from a database comprising fully sequenced proteomes and generated a cluster map based on their all-against-all pairwise sequence similarity. Our results show that the hemerythrin-like superfamily comprises a large number of protein families which can be classified into three broad groups on the basis of their cation-coordinating residues: (a) signal-transduction and oxygen-carrier hemerythrins (H-HxxxE-HxxxH-HxxxxD); (b) hemerythrin-like (H-HxxxE-H-HxxxE); and, (c) metazoan F-box proteins (H-HExxE-H-HxxxE). Interestingly, all but two hemerythrin-like families exhibit internal sequence and structural symmetry, suggesting that a duplication event may have led to the origin of the hemerythrin domain.

The absence of the hydrophobic norvaline and norleucine in the inventory of protein amino acids is readdressed. The well-documented intracellular accumulation of these two amino acids results from the low-substrate specificity of the branched-chain amino acid biosynthetic enzymes that act over a number of related α-ketoacids. The lack of absolute substrate specificity of leucyl-tRNA synthase leads to a mischarged norvalyl-tRNA(Leu) that evades the translational proofreading activities and produces norvaline-containing proteins, (cf. Apostol et al. J Biol Chem 272:28980-28988, 1997). A similar situation explains the presence of minute but detectable amounts of norleucine in place of methionine. Since with few exceptions both leucine and methionine are rarely found in the catalytic sites of most enzymes, their substitution by norvaline and norleucine, respectively, would have not been strongly hindered in small structurally simple catalytic polypeptides during the early stages of biological evolution. The report that down-shifts of free oxygen lead to high levels of intracellular accumulation of pyruvate and the subsequent biosynthesis of norvaline (Soini et al. Microb Cell Factories 7:30, 2008) demonstrates the biochemical and metabolic consequences of the development of a highly oxidizing environment. The results discussed here also suggest that a broader definition of biomarkers in the search for extraterrestrial life may be required.

BackgroundThe evolution of oxygenic photosynthesis during Precambrian times entailed the diversification of strategies minimizing reactive oxygen species-associated damage. Four families of oxygen-carrier proteins (hemoglobin, hemerythrin and the two non-homologous families of arthropodan and molluscan hemocyanins) are known to have evolved independently the capacity to bind oxygen reversibly, providing cells with strategies to cope with the evolutionary pressure of oxygen accumulation. Oxygen-binding hemerythrin was first studied in marine invertebrates but further research has made it clear that it is present in the three domains of life, strongly suggesting that its origin predated the emergence of eukaryotes.ResultsOxygen-binding hemerythrins are a monophyletic sub-group of the hemerythrin/HHE (histidine, histidine, glutamic acid) cation-binding domain. Oxygen-binding hemerythrin homologs were unambiguously identified in 367/2236 bacterial, 21/150 archaeal and 4/135 eukaryotic genomes. Overall, oxygen-binding hemerythrin homologues were found in the same proportion as single-domain and as long protein sequences. The associated functions of protein domains in long hemerythrin sequences can be classified in three major groups: signal transduction, phosphorelay response regulation, and protein binding. This suggests that in many organisms the reversible oxygen-binding capacity was incorporated in signaling pathways. A maximum-likelihood tree of oxygen-binding hemerythrin homologues revealed a complex evolutionary history in which lateral gene transfer, duplications and gene losses appear to have played an important role.ConclusionsHemerythrin is an ancient protein domain with a complex evolutionary history. The distinctive iron-binding coordination site of oxygen-binding hemerythrins evolved first in prokaryotes, very likely prior to the divergence of Firmicutes and Proteobacteria, and spread into many bacterial, archaeal and eukaryotic species. The later evolution of the oxygen-binding hemerythrin domain in both prokaryotes and eukaryotes led to a wide variety of functions, ranging from protection against oxidative damage in anaerobic and microaerophilic organisms, to oxygen supplying to particular enzymes and pathways in aerobic and facultative species.

Mechanisms of emergence and divergence of protein folds pose central questions in biological sciences. Incremental mutation and stepwise adaptation explain relationships between topologically similar protein folds. However, the universe of folds is diverse and riotous, suggesting more potent and creative forces are at play. Sequence and structure similarity are observed between distinct folds, indicating that proteins with distinct folds may share common ancestry. We found evidence of common ancestry between three distinct β-barrel folds: Scr kinase family homology (SH3), oligonucleotide/oligosaccharide-binding (OB), and cradle loop barrel (CLB). The data suggest a mechanism of fold evolution that interconverts SH3, OB, and CLB. This mechanism, which we call creative destruction, can be generalized to explain many examples of fold evolution including circular permutation. In creative destruction, an open reading frame duplicates or otherwise merges with another to produce a fused polypeptide. A merger forces two ancestral domains into a new sequence and spatial context. The fused polypeptide can explore folding landscapes that are inaccessible to either of the independent ancestral domains. However, the folding landscapes of the fused polypeptide are not fully independent of those of the ancestral domains. Creative destruction is thus partially conservative; a daughter fold inherits some motifs from ancestral folds. After merger and refolding, adaptive processes such as mutation and loss of extraneous segments optimize the new daughter fold. This model has application in disease states characterized by genetic instability. Fused proteins observed in cancer cells are likely to experience remodeled folding landscapes and realize altered folds, conferring new or altered functions.