[Show abstract][Hide abstract] ABSTRACT: Nanobiomotors perform various important functions in the cell, and they also emerge as potential vehicle for drug delivery. These proteins employ conserved ATPase domains to convert chemical energy to mechanical work and motion. Several archaeal nucleic acid nanobiomotors, such as DNA helicases that unwind double-stranded DNA molecules during DNA damage repair, have been characterized in details. XPB, XPD and Hjm are SF2 family helicases, each of which employs two ATPase domains for ATP binding and hydrolysis to drive DNA unwinding. They also carry additional specific domains for substrate binding and regulation. Another helicase, HerA, forms a hexameric ring that may act as a DNA-pumping enzyme at the end processing of double-stranded DNA breaks. Common for all these nanobiomotors is that they contain ATPase domain that adopts RecA fold structure. This structure is characteristic for RecA/RadA family proteins and has been studied in great details. Here we review the structural analyses of these archaeal nucleic acid biomotors and the molecular mechanisms of how ATP binding and hydrolysis promote the conformation change that drives mechanical motion. The application potential of archaeal nanobiomotors in drug delivery has been discussed.
[Show abstract][Hide abstract] ABSTRACT: A putative protease gene (tldD) was identified from studying tolerance of letD encoding CcdB toxin of a toxin-antidote system of the F plasmid in Escherichia coli. While this gene is evolutionarily conserved in Archaea and Bacrteria, proteolytic activity of encoded proteins remained to be demonstrated experimentally. Here we studied Sso0660, an archaeal TldD homologue encoded in Sulfolobus solfataricus by over-expression of the recombinant protein and characterisation of the purified enzyme. We found that the enzyme is active in degrading azocasein and FTC-BSA substrates. Protease inhibitor studies showed that EDTA and ο-phenanthroline, two well-known metalloprotease inhibitors, either abolished completely or strongly inhibited the enzyme activity and flame spectrometric analysis showed zinc ion is a co-factor of the protease. Furthermore, the protein forms disulphide bond via the C416 residue, yielding protein dimer that is the active form of the enzyme. These results establish for the first time that TldDs encode zinc-containing proteases, classifying them as a family in the metalloprotease class.
[Show abstract][Hide abstract] ABSTRACT: Rad51/RadA paralogs found in eukaryotes and euryarchaea play important roles during recombination and repair, and mutations in one of the human Rad51 paralogs, Rad51C, are associated with breast and ovarian cancers. The hyperthermophilic crenarchaeon Sulfolobus tokodaii encodes four putative RadA paralogs and studies on these proteins may assist in understanding the functions of human Rad51 paralogs. Here, we report the biochemical characterization of stRadC2, a S. tokodaii RadA paralog. Pull-down assays revealed that the protein was able to interact with the recombinase, RadA, and the Holliday junction endonuclease, Hjc. stRadC2 inhibited the strand exchange activity of RadA and facilitated Hjc-mediated Holliday junction DNA cleavage in vitro. RT-PCR analysis revealed that stRadC2 transcription was immediately reduced after UV irradiation, but was restored to normal levels at the late stages of DNA repair. Our results suggest that stRadC2 may act as an anti-recombination factor in DNA recombinational repair in S. tokodaii.
Science China. Life sciences 03/2012; 55(3):261-7. · 2.02 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: XPB helicase is the largest subunit of transcription factor IIH (TFIIH), a ten-subunit protein complex essential for transcription initiation and nucleotide excision repair (NER) in Eukarya. Two XPB homologues (XPBI and XPBII) are present in the genome of most crenarchaeota, one of the two major phyla of archaea; however, the biochemical properties have not been fully characterized and their cellular roles have not been clearly defined. Here, we report that XPBI from the hyperthermophilic crenarchaeon Sulfolobus tokodaii (StoXPBI) is able to destabilize double-stranded DNA (dsDNA) helix independent of ATP (designated as dsDNA melting activity). This activity is inhibited by single-stranded DNA (ssDNA) and relies on the unique N-terminal domain of StoXPBI, which is also likely responsible for the intrinsic strong ssDNA binding activity of StoXPBI as revealed by deletion analysis. We demonstrate that the ATPase activity of StoXPBII is remarkably stimulated by StoBax1, a nuclease partner of StoXPBII. The role of the unique dsDNA melting activity of XPBI in NER in archaea was discussed.