A novel type of uracil-DNA glycosylase mediating repair of hydrolytic DNA damage in the extremely thermophilic eubacterium Thermus thermophilus.
Spontaneous hydrolytic deamination of DNA cytosine and 5-methyl-cytosine residues is an abundant source of C/G (5-meC/G) to T/A transition mutations. As a result of this pressure, at least six different families of enzymes have evolved that initiate repair at U/G (T/G) mispairs, the relevant pre-mutagenic intermediates. The necessarily higher rate of the process at elevated temperatures must pose a correspondingly accentuated problem to contemporary thermophilic organisms and may have been a serious bottleneck in early evolution when life passed through a phase of very high ambient temperatures. Here we show that Thermus thermophilus, an aerobic, Gram-negative eubacterium thriving at up to 85 degrees C, harbors two uracil-DNA glycosylases (UDGs), termed TTUDGA and TTUDGB. According to both amino acid sequence and enzymatic properties, TTUDGA clearly belongs to the family of 'thermostable UDGs'. TTUDGB shares with TTUDGA 23% sequence identity, but differs from it in profound functional aspects. TTUDGB, unlike TTUDGA, does not act upon uracil residues in the context of single-stranded DNA whereas both enzymes process various double-stranded substrates, albeit with different preferences. TTUDGB shows a number of sequence features characteristic of the UDG superfamily, but surprisingly lacks any polar residue within its so-called motif 1 (GLAPG-X(10)-F). This finding is in conflict with a previously assumed crucial catalytic role of motif 1 in water activation and supports a more recently suggested alternative of a dissociative ('S(N)1-type') reaction mechanism. Together, the characteristics of TTUDGB and its homologs in other organisms define a novel family of UDG repair enzymes.
Starkuviene
Vytaute
V
Abteilung Molekulare Genetik und Präparative Molekularbiologie and Göttingen Genomics Laboratory, Institut für Mikrobiologie und Genetik, Georg-August-Universität Göttingen, Grisebachstrasse 8, D-37077 Göttingen, Germany.
Fritz
Hans-Joachim
HJ
D020816:Amino Acid Motifs / Q000235:genetics; D000595:Amino Acid Sequence; D019295:Computational Biology; D004249:DNA Damage; D045647:DNA Glycosylases; D004260:DNA Repair; D019641:DNA, Archaeal / Q000235:genetics / Q000378:metabolism*; D016680:Genome, Bacterial; D007527:Isoenzymes / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D009699:N-Glycosyl Hydrolases / Q000235:genetics / Q000378:metabolism*; D010802:Phylogeny; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D016963:Thermus thermophilus / Q000201:enzymology* / Q000235:genetics; D051981:Uracil-DNA Glycosidase
D019641:DNA, Archaeal; D007527:Isoenzymes; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; D051981:Uracil-DNA Glycosidase