An unprecedented nucleic acid capture mechanism for excision of DNA damage

Abstract

DNA glycosylases that remove alkylated and deaminated purine nucleobases are essential DNA repair enzymes that protect the genome, and at the same time confound cancer alkylation therapy, by excising cytotoxic N3-methyladenine bases formed by DNA targeting anticancer compounds. The basis for glycosylase specificity toward N3- and N7-alkylpurines is believed to result from intrinsic instability of the modified bases and not from direct enzyme functional group chemistry. Here, we present crystal structures of the recently discovered Bacillus cereus AlkD glycosylase in complex with DNAs containing alkylated, mismatched, and abasic nucleotides. Unlike other glycosylases, AlkD captures the extrahelical lesion in a solvent-exposed orientation, providing the first illustration for how hydrolysis of N3- and N7-alkylated bases may be facilitated by increased lifetime out of the DNA helix. The structures and supporting biochemical analysis of base flipping and catalysis reveal how AlkD’s HEAT-repeats distort the DNA backbone to detect non-Watson-Crick base pairs without duplex intercalation.

Structure of a DNA glycosylase that unhooks interstrand cross-links

“…Recently, the AlkZ protein was shown to act on AZB ICLs and monoadducts through hydrolysis of one or more of the N-glycosidic bonds (16), but whether the enzyme would act on adducts other than AZB was unclear. DNA glycosylases that excise alkylated DNA lesions often exhibit activity for a number of N3-and N7-methylated purines in addition to their preferred substrates (21,22). For example, Bacillus cereus AlkD displays robust activity for bulky minor groove N3-yatakemycinyldeoxyadenosine lesions, but also cleaves N3-methyldeoxyadenosine and N7-methyldeoxyguanosine (d7mG) (23,24).…”

“…The bacterial AlkD glycosylase is able to recognize and cleave deoxyadenosine adducts of the bulky natural product yatakemycin (YTM) without rotating the lesion from the duplex (24). Like AlkZ, AlkD adopts a C-shaped fold that engages DNA along the concave surface (22). The AlkD active site positions a catalytic aspartate and two catalytic tryptophan side chains (25) against the deoxyribose of the alkylated adenosine, which is displaced only slightly into the minor groove of the DNA.…”

Thermally Activated Delayed Fluorescence Organic Dots (TADF Odots) for Time‐Resolved and Confocal Fluorescence Imaging in Living Cells and In Vivo

“…Fluorescence imaging has been demonstrated as an indispensable and versatile tool in tracking and understanding complex biological environments both in vivo and in vitro 1, 2. Compared to other imaging modalities, fluorescence imaging offers high spatiotemporal resolution as a noninvasive technique 1, 2, 3.…”

“…Compared to other imaging modalities, fluorescence imaging offers high spatiotemporal resolution as a noninvasive technique 1, 2, 3. It can probe the deeper layers of a specimen at ambient conditions and also enable spectroscopic diagnosis with chemical sensitivity 4.…”

Arabidopsis thaliana Tic110, involved in chloroplast protein translocation, contains at least fourteen highly divergent heat-like repeated motifs

“…Up to now, Tic110 is considered to be of eukaryotic origin (Reumann & Keegstra 1999;Kalanon & McFadden 2008;Kovács-Bogdán et al 2010). However, HEAT repeats are well distributed in many prokaryotic proteins, for example, Interpro database (Hunter et al 2009) registers 840 bacteria and 173 cyanobacteria HEAT proteins (entry IPR000357) and structures are available in the PDB (Rubinson et al 2008(Rubinson et al , 2010. Because prokaryotes were the first organisms to evolve (Vellai & Vida 1999), it is reasonable to suggest that Tic110 might have been newly developed in the arising plant cell in concert with chloroplast development, i.e., a dual origin or lost from ancestral cyanobacteria (Reumann & Keegstra 1999;Reumann et al 2005).…”