Conserved structural motifs governing the stoichiometric repair of alkylated DNA by O6-alkylguanine-DNA alkyltransferase

Direct Reversal of DNA Alkylation Damage

“…103 These four residues are conserved in all mammalian sequences of the protein but are missing from both S. cerevisiae and bacterial homologues. 104 Further biochemical studies showed that although the zinc(II) site is far from the reaction center, binding a zinc(II) ion serves to lower the pK a of the Cys145 even further than that is in the apo protein, which increases the reactivity of the protein. 105,106 A detailed Cys145 activation and repair mechanism was proposed based on the stcuture.…”

Recent progress on the Ada response for inducible repair of DNA alkylation damage

“…The activated Ada protein induces expression of several genes resulting in increased DNA repair and probable destruction of certain alkylating agents methyltransferase (AGT) also implicates a HTH motif in DNA binding and an 'arginine finger' to extrude the O 6 -alkG nucleotide from the duplex DNA. The extrahelical base can then reach the recessed AGT active site (Daniels and Tainer, 2000). These conserved motifs in the E. coli C-Ada19 domain are shown in Figure 3.…”

“…Within the proposed HTH of the C-terminal domain is a conserved 'arginine finger' which is implicated in nucleotide flipping. An invariant Asn-hinge couples the recognition helix (second helix) to the active site cysteine in this domain (Daniels and Tainer, 2000) Eukaryotic organisms do not appear to have a DNA methyltransferase that repairs methylphosphotriesters. Presumably this is not required as eukaryotes do not have an inducible response equivalent to that in bacteria.…”

Structural studies of MJ1529, an O6-methylguanine-DNA methyltransferase

“…In both MJ1529 and the crystal structures, R111 in helix f is exposed and in a position to interact with the negatively charged backbone of DNA. It is proposed that this invariant residue acts as an "arginine finger," injecting itself into the base stack and promoting the flipping out of the guanine base, and stabilizing the resulting structure, akin to other DNA repair enzymes that use amino acids to promote base-flipping 21,[40][41][42] . A112 is proposed to make sequence-independent hydrophobic contacts with the DNA 21,31,38 .…”

“…One interaction that could not be detected (not likely due to its absence but due to data limitations) is a hydrogen bond between the sidechains of invariant N120 and C128. It is this interaction that is broken by alkylation, and results in a subtle structural change that destabilizes the protein, and in the case of hMGMT, signals the protein for ubiquitin dependent degradation 40 .…”