Proteins’ interactions with ancient ligands may reveal how molecular recognition emerged and evolved. We explore how proteins recognize adenine: a planar rigid fragment found in the most common and ancient ligands. We have developed a computational pipeline that extracts protein–adenine complexes from the Protein Data Bank, structurally superimposes their adenine fragments, and detects the hydrogen bonds mediating the interaction. Our analysis extends the known motifs of protein–adenine interactions in the Watson–Crick edge of adenine and shows that all of adenine’s edges may contribute to molecular recognition. We further show that, on the proteins' side, binding is often mediated by specific amino acid segments (“themes”) that recur across different proteins, such that different proteins use the same themes when binding the same adenine-containing ligands. We identify numerous proteins that feature these themes and are thus likely to bind adenine-containing ligands. Our analysis suggests that adenine binding has emerged multiple times in evolution.

Outer-membrane beta barrels (OMBBs) are found in the outer membrane of gram-negative bacteria and eukaryotic organelles. OMBBs fold as antiparallel β-sheets that close onto themselves, forming pores that traverse the membrane. Currently known structures include only one barrel, of 8 to 36 strands, per chain. The lack of multi-OMBB chains is surprising, as most OMBBs form oligomers, and some function only in this state. Using a combination of sensitive sequence comparison methods and coevolutionary analysis tools, we identify many proteins combining multiple beta barrels within a single chain; combinations that include eight-stranded barrels prevail. These multibarrels seem to be the result of independent, lineage-specific fusion and amplification events. The absence of multibarrels that are universally conserved in bacteria with an outer membrane, coupled with their frequent de novo genesis, suggests that their functions are not essential but rather beneficial in specific environments. Adjacent barrels of complementary function within the same chain may allow for functions beyond those of the individual barrels.

Outer membrane beta barrels (OMBBs) are found in the outer membrane of Gram-negative bacteria and eukaryotic organelles. OMBBs fold as antiparallel β-sheets that close onto themselves, forming pores that traverse the membrane. Currently known structures include only one barrel, of 8-36 strands, per chain. The lack of multi-OMBB chains is surprising, as most OMBBs form oligomers and some function only in this state. Using a combination of sensitive sequence-comparison methods and co-evolutionary analysis tools, we identify many proteins combining multiple beta barrels within a single chain; combinations that include 8-stranded barrels prevail. These multi-barrels seem to be the result of independent, lineage-specific fusion and amplification events. The absence of multi-barrels that are universally conserved in bacteria with an outer membrane, coupled with their frequent de novo genesis suggests that their functions are not essential, but rather beneficial in specific environments. Adjacent barrels of complementary function within the same chain may allow for new functions beyond those of the individual barrels.

Currently known G protein-coupled receptors (GPCRs) have a single transmembrane domain. Many GPCRs form dimers that have two transmembrane domains (one per protein), and there are indications that this dimeric interaction is functionally meaningful. Here, based on sequence analysis and structure predictions, we report the existence of 57 proteins with two, three, or four GPCR domains within the same protein chain. We analyze the structures of these multi-GPCRs and show that almost all have DRY/NPxxY motifs, a strong indication of signaling activity. By homology, most of the multi-GPCRs that we identified are olfactory-related; a few are chemokine-related. Multi-GPCR candidates are found in various Chordata species including fish, camel, marmite, Chinese hamster, and new world monkeys. The discovery of receptors with multiple transmembrane domains suggests the possibility for signal regulation and amplification within an individual receptor, revealing another step in GPCR evolution and a new layer of complexity in signal transduction.

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