ContentsI. Introduction 1221 II. Types of Base-Excision Repair Glycosylases 1223 A. Uracil-DNA Glycosylase (UDG) 1223 B. Mismatch-Specific Thymine-DNA Glycosylase (TDG) and Related Double-Strand Specific Uracil DNA Glycosylase (dsUDG) 1226 C. Alkylated Base Removal 1227 D. Endonuclease III and Related Enzymes 1229 E. Pyrimidine Dimer Glycosylases 1230 F. The FPG Protein (Fapy Glycosylase or MutM) 1231

The DNA repair enzyme MutY plays an important role in the prevention of DNA mutations caused by the oxidatively damaged lesion 7,8-dihydro-8-oxo-2′-deoxyguanosine (OG) by removal of misincorporated adenine residues in OG:A mismatched base pairs using N-glycosylase activity. MutY also has glycosylase activity toward adenine in the mismatched base-pairs G:A and C:A. We have investigated the interaction of MutY with DNA duplexes containing the 2′-deoxyadenosine (A) analogs 2′-deoxytubercidin (7-deaza-2′-deoxyadenosine, Z) and 2′-deoxyformycin A (F). Both F and Z should effectively mimic the recognition properties of A but be resistant to the glycosylase activity of MutY, owing to their structural properties. Thus, these derivatives will provide a method for forming a stable MutY-substrate analog complex amenable to structural and biochemical investigation. We find that oligonucleotide duplexes containing OG/G:F and OG/G:Z base-pairs are not substrates for MutY as expected. Using a gel retardation method to measure relevant K d values, we determined that MutY has an increased association with duplexes containing OG/G:F and OG/G:Z base-pairs over their OG/G:C counterparts. Interestingly, MutY has a higher affinity for the F-containing duplexes than the Z counterparts. Additionally, MutY binds to the OG:F and G:F duplexes with a similar, albeit lower, affinity as the substrate OG:A and G:A duplexes. In footprinting experiments using methidiumpropyl-EDTA-Fe(II), a region of the duplex surrounding the OG:F base-pair is observed which is protected by MutY from hydroxyl radical cleavage. These results provide additional evidence for specific recognition of the OG:F base-pair within the DNA duplex. Furthermore, these results also illustrate the utility of OG:F duplexes for providing information regarding the MutY-mismatched DNA complex which could not be obtained with the normal OG:A substrate since a footprint on both strands of the duplex could only be observed with the OG:Fcontaining duplex. These substrate analog duplexes will provide avenues for structural analysis of the MutYmismatched DNA complex and for investigating the properties of the unusual [4Fe-4S] center in MutY.

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We present the results from a photometric monitoring program of 15 open cluster stars and one weak-lined T Tauri star during late 1993/early 1994. Several slow rotators which are members of the Alpha Persei, Pleiades, and Hyades open clusters have been monitored and period estimates derived. Using all available Pleiades stars with photometric periods together with current X-ray flux measurements, we illustrate the X-ray activity/rotation relation among Pleiades late-G/K dwarfs. The data show a clear break in the rotation-activity relation around P-6-1 days-in general accordance with previous results using more heterogeneous samples of G/K stars.

The Escherichia coli adenine glycosylase MutY is involved in the repair of 7,8-dihydro-8-oxo-2'-deoxyguanosine (OG):A and G:A mispairs in DNA. DNA strand cleavage via beta-elimination (beta-lyase) activity coupled with MutY's removal of misincorporated adenine bases was sought using both qualitative and quantitative methods. The qualitative assays demonstrate formation of a Schiff base intermediate which is characteristic of DNA glycosylases catalyzing a concomitant beta-lyase reaction. Borohydride reduction of the Schiff base results in the formation of a covalent DNA-MutY adduct which is easily detected in SDS-PAGE experiments. However, quantitative activity assays which monitor DNA strand scission accompanying base release suggest MutY behaves as a simple monofunctional glycosylase. Treatment with base effects DNA strand cleavage at apurinic/apyrimidinic (AP) sites arising via simple glycosylase activity. The amount of cleaved DNA in MutY reactions treated with base is much greater than that in non-base treated reactions, indicating that AP site generation by MutY is not associated with a concomitant beta-lyase step. As standards, identical assays were performed with a known monofunctional enzyme (uracil DNA glycosylase) and a known bifunctional glycosylase/lyase (FPG), the results of which were used in comparison with those of the MutY experiments. The apparent inconsistency between the data obtained for MutY by the qualitative and quantitative methods underscores the current debate surrounding the catalytic activity of this enzyme, and a detailed explanation of this controversy is proposed. The work presented here lays ground for the identification of specific active site residues responsible for the chemical mechanism of MutY enzyme catalysis.