CC1, a Novel Crenarchaeal DNA Binding Protein

Abstract

The genomes of the related crenarchaea Pyrobaculum aerophilum and Thermoproteus tenax lack any obvious gene encoding a single-stranded DNA binding protein (SSB). SSBs are essential for DNA replication, recombination, and repair and are found in all other genomes across the three domains of life. These two archaeal genomes also have only one identifiable gene encoding a chromatin protein (the Alba protein), while most other archaea have at least two different abundant chromatin proteins. We performed a biochemical screen for novel nucleic acid binding proteins present in cell extracts of T. tenax. An assay for proteins capable of binding to a single-stranded DNA oligonucleotide resulted in identification of three proteins. The first protein, Alba, has been shown previously to bind single-stranded DNA as well as duplex DNA. The two other proteins, which we designated CC1 (for crenarchaeal chromatin protein 1), are very closely related to one another, and homologs are restricted to the P. aerophilum and Aeropyrum pernix genomes. CC1 is a 6-kDa, monomeric, basic protein that is expressed at a high level in T. tenax. This protein binds single-and double-stranded DNAs with similar affinities. These properties are consistent with a role for CC1 as a crenarchaeal chromatin protein.

Displacement of the canonical single-stranded DNA-binding protein in the Thermoproteales

“…OB fold-containing SSB proteins have been detected in all three domains of life, but, as we reported previously, one group of Crenarchaea, the Thermoproteales, appear to lack an identifiable SSB-encoding gene (25). There are now 10 fully sequenced genomes in this group (Thermoproteus tenax, Thermoproteus uzoniensis, Thermoproteus neutrophilius, Caldivirga maquilingensis, Pyrobaculum aerophilum, Pyrobaculum arsenaticum, Pyrobaculum islandicum, Pyrobaculum calidifontis, Vulcanisaeta moutnovskia, and Vulcanisaeta distributa) that lack any identifiable ssb genes.…”

Heterologous Expression of the Alba Protein from the Hyperthermophilic Archaeon Aeropyrum Pernix

“…Results of the protein/ DNA binding obtained were similar to those seen for other Alba/DNA gel mobility shift assays, where DNA delaying was seen. 6,9,10,14,22 There was a delaying effect of Alba1 binding to the DNA, followed by a precipitating effect when the critical DNA bp:protein molar ratio was exceeded (> 2:1). For Alba2, precipitation of DNA (> 10:1) was observed primarily.…”

“…7 kDa small basic proteins). [6][7][8][9][10] Two Alba protein sequences were identified in the A. pernix genome, APE1832.1 (Alba1) and APE1823 (Alba2), which appear to originate from gene duplication. 11 The Alba1 protein consists of 94 amino acid residues (10.3 kDa; pI = 9.5), while Alba2 consists of 102 amino acid residues (11.4 kDa, pI = 9.1).…”

Role of RPA Proteins in Radiation Repair and Recovery

“…To our knowledge, there have been no reports of genomes lacking an RPA/SSB homolog, with the possible exception of the crenarchaea Pyrobaculum aerophilum and Aeropyrum pernix (Luo, et al, 2007). There are numerous excellent and extensive reviews covering the history, genetics and biochemical characterization of E. coli SSB and its encoding gene (Lohman & Ferrari, 1994;Meyer & Laine, 1990;Shereda, et al, 2008).…”

“…Despite the similarity to bacterial SSB, and the ability of the gene to complement the lethality of an E. coli ssb mutation, the DNA-binding domain of this protein more closely resembles those of eukaryotic RPA1 (Haseltine & Kowalczykowski, 2002;Kerr, et al, 2003). Although the genomes of two related crenarchaea, P. aerophilum and A. pernix, have been reported to contain no obvious RPA or SSB homolog (Luo, et al, 2007), a previous study found a Sulfolobus-like SSB in A. pernix (Haseltine & Kowalczykowski, 2002). Given the recent identification of a novel SSB in D. radiodurans, and the fundamental role that ssDNA binding proteins play in replication and repair, it is likely that a protein carrying out these essential functions will be found in all organisms.…”