Inteins at Eleven Distinct Insertion Sites in Archaeal Helicase Subunit MCM Exhibit Varied Architectures and Activity Levels Across Archaeal Groups

Background/Objectives: Inteins are mobile genetic elements invading highly conserved genes across all domains of life and viruses. Five active intein insertion sites (MCM-a through e) have been identified and studied in the archaeal replicative helicase gene mcm, making the MCM protein an ideal system for dissecting the dynamics of multi-intein genes. However, work in this system thus far has been limited to particular archaeal groups. To better understand the dynamics and diversity of these inteins, MCM homologs spanning all archaeal groups were extracted from NCBI’s non-redundant protein sequence database, and the distribution and structural architectures of their inteins were thoroughly characterized. Methods: The amino acid sequences of 4,243 archaeal MCM homologs were retrieved from NCBI’s non-redundant protein sequence database. These sequences were systematically assessed for their intein content through within-group multiple sequence alignments. To characterize the inteins present at each site, extensive intein structure predictions and comparisons were performed. Phylogenetic analyses were used to investigate intein relatedness between and within sites, as well as the distribution of different MCM inteins in geographically overlapping populations of archaea. Results: In total, 11 active MCM intein insertion sites were identified, expanding on the previously known five. The insertion sites have varied invasion activity levels across archaeal groups, with Nanobdellati (DPANN) being the only group with all 11 sites active. In all but two (Methanonatronarchaeia and Hadarchaeota) of the archaeal groups studied where inteins were present, there was at least one case of an MCM homolog invaded by more than one intein. With respect to intein structure, within-intein insertions bearing semblance to DNA-binding domains were identified, with varied presence between MCM inteins. Additionally, a study of archaeal MCM sequences of samples collected from the Atacama Desert in June 2013 revealed high MCM intein diversity levels. Conclusions: We present six new active intein insertion sites in archaeal MCM, expanding on the five previously known sites. No individual contained more than four MCM inteins simultaneously. Many inteins analyzed contained insertions bearing similarity to DNA-binding helix-turn-helix domains suggesting potential involvement in the intein homing process. Additionally, the high levels of MCM intein diversity observed in archaea from the Atacama Desert provide strong support for a co-existence model of intein persistence.