Role of the basic amino acid cluster and Glu-23 in pyrimidine dimer glycosylase activity of T4 endonuclease V.

Doi, Tomoko; Recktenwald, A.; Karaki, Yoko; Kikuchi, Masakazu; Morikawa, Kosuke; Ikehara, Morio; Inaoka, Tetsuya; Hori, Naoko; Ohtsuka, Eiko

doi:10.1073/pnas.89.20.9420

Cited by 52 publications

(64 citation statements)

References 15 publications

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“…Two amino acids critical for T4-PDG catalysis are known from x-ray crystal (Morikawa et al 1992) and cocrystal (Vassylyev et al 1995) structures and studies of site-directed mutations (Hori et al 1992;Doi et al 1992). These amino acids are Thr 2 (Dodson et al 1993;Schrock and Lloyd 1991) and Glu 23 (Hori et al 1992;Doi et al 1992;Manuel et al 1995).…”

Section: Discussionmentioning

confidence: 99%

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Intron Conservation in a UV-Specific DNA Repair Gene Encoded by Chlorella Viruses

Sun

Liu²,

McCullough

et al. 2000

J Mol Evol

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Abstract. Large dsDNA-containing chlorella viruses encode a pyrimidine dimer-specific glycosylase (PDG) that initiates repair of UV-induced pyrimidine dimers. The PDG enzyme is a homologue of the bacteriophage T4-encoded endonuclease V. The pdg gene was cloned and sequenced from 42 chlorella viruses isolated over a 12-year period from diverse geographic regions. Surprisingly, the pdg gene from 15 of these 42 viruses contain a 98-nucleotide intron that is 100% conserved among the viruses and another 4 viruses contain an 81-nucleotide intron, in the same position, that is nearly 100% identical (one virus differed by one base). In contrast, the nucleotides in the pdg coding regions (exons) from the introncontaining viruses are 84 to 100% identical. The introns in the pdg gene have 5Ј-AG/GTATGT and 3Ј-TTGCAG/ AA splice site sequences which are characteristic of nuclear-located, spliceosomal processed pre-mRNA introns. The 100% identity of the 98-nucleotide intron sequence in the 15 viruses and the near-perfect identity of an 81-nucleotide intron sequence in another 4 viruses imply strong selective pressure to maintain the DNA sequence of the intron when it is in the pdg gene. However, the ability of intron-plus and intron-minus viruses to repair UV-damaged DNA in the dark was nearly identical. These findings contradict the widely accepted dogma that intron sequences are more variable than exon sequences.

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Section: Discussionmentioning

confidence: 99%

“…These amino acids are Thr 2 (Dodson et al 1993;Schrock and Lloyd 1991) and Glu 23 (Hori et al 1992;Doi et al 1992;Manuel et al 1995). These two residues are integral for labilizing the glycosyl bond of the 5Ј pyrimidine of the dimer and catalyzing phosphodiester backbone cleavage by ␤-elimination.…”

Section: Discussionmentioning

confidence: 99%

Intron Conservation in a UV-Specific DNA Repair Gene Encoded by Chlorella Viruses

Sun

Liu²,

McCullough

et al. 2000

J Mol Evol

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“…residue that is known to dramatically reduce activity in homologs (Doi et al, 1992;Goosen and Moolenaar, 2008), suggesting a diminished ability to repair UV-damaged DNA in eSS120 and eMIT9313 ecotypes. Indeed, the high-light adapted strain MED4, which encodes photolyase, has a greater tolerance to UV exposure than low-light adapted strain MIT9313 (Osburne et al, 2010), which lacks photolyase but encodes pyrimidine dimmer glycosylase.…”

Section: Response To Light Shock In Cultured Isolatesmentioning

confidence: 99%

Temporal dynamics of Prochlorococcus ecotypes in the Atlantic and Pacific oceans

et al. 2010

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To better understand the temporal and spatial dynamics of Prochlorococcus populations, and how these populations co-vary with the physical environment, we followed monthly changes in the abundance of five ecotypes-two high-light adapted and three low-light adapted-over a 5-year period in coordination with the Bermuda Atlantic Time Series (BATS) and Hawaii Ocean Time-series (HOT) programs. Ecotype abundance displayed weak seasonal fluctuations at HOT and strong seasonal fluctuations at BATS. Furthermore, stable 'layered' depth distributions, where different Prochlorococcus ecotypes reached maximum abundance at different depths, were maintained consistently for 5 years at HOT. Layered distributions were also observed at BATS, although winter deep mixing events disrupted these patterns each year and produced large variations in ecotype abundance. Interestingly, the layered ecotype distributions were regularly reestablished each year after deep mixing subsided at BATS. In addition, Prochlorococcus ecotypes each responded differently to the strong seasonal changes in light, temperature and mixing at BATS, resulting in a reproducible annual succession of ecotype blooms. Patterns of ecotype abundance, in combination with physiological assays of cultured isolates, confirmed that the low-light adapted eNATL could be distinguished from other low-light adapted ecotypes based on its ability to withstand temporary exposure to high-intensity light, a characteristic stress of the surface mixed layer. Finally, total Prochlorococcus and Synechococcus dynamics were compared with similar time series data collected a decade earlier at each location. The two data sets were remarkably similar-testimony to the resilience of these complex dynamic systems on decadal time scales.

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“…This glycosylase also possesses a concomitant AP lyase activity, resulting in the production of a 3Ј ␣,␤-unsaturated aldehyde and a 5Ј-phosphate via a ␤-elimination reaction at the glycosylase generated abasic (AP) site. Biochemical and structural analyses have demonstrated the involvement of two key residues in the catalytic activity of the enzyme, Glu-23 and the N-terminal Thr-2 (12)(13)(14)(15)(16)(17)(18).…”

mentioning

confidence: 99%

The Role of Base Flipping in Damage Recognition and Catalysis by T4 Endonuclease V

McCullough

Dodson

Schärer

et al. 1997

Journal of Biological Chemistry

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The process of moving a DNA base extrahelical (base flipping) has been shown in the co-crystal structure of a UV-induced pyrimidine dimer-specific glycosylase, T4 endonuclease V, with its substrate DNA. Compared with other enzymes known to use base flipping, endonuclease V is unique in that it moves the base opposite the target site extrahelical, rather than moving the target base itself. Utilizing substrate analogs and catalytically inactive mutants of T4 endonuclease V, this study investigates the discrete steps involved in damage recognition by this DNA repair enzyme. Specifically, fluorescence spectroscopy analysis shows that fluorescence changes attributable to base flipping are specific for only the base directly opposite either abasic site analogs or the 5 -thymine of a pyrimidine dimer, and no changes are detected if the 2-aminopurine is moved opposite the 3 -thymine of the pyrimidine dimer. Interestingly, base flipping is not detectable with every specific binding event suggesting that damage recognition can be achieved without base flipping. Thus, base flipping does not add to the stability of the specific enzyme-DNA complex but rather induces a conformational change to facilitate catalysis at the appropriate target site. When used in conjunction with structural information, these types of analyses can yield detailed mechanistic models and critical amino acid residues for extrahelical base movement as a mode of damage recognition.The initiating events in base excision repair are performed by a class of enzymes, DNA glycosylases. These damage-specific enzymes are responsible for recognizing and binding to damaged bases and catalyzing the cleavage of the N-C 1 Ј glycosydic bond linking the damaged base to the sugar phosphate backbone. Glycosylases thus provide the specificity to base excision repair. The precise mechanism by which these enzymes discriminate between nontarget and target DNA bases is beginning to be elucidated for a few glycosylases due to insights from high resolution x-ray crystallographic structures (reviewed in Ref. 1). In particular, uracil DNA glycosylases and a catalytically inactive mutant of T4 endonuclease V have been co-crystallized with their product or substrate DNAs, respectively, and shown to move a base extrahelical (nucleotide), placing the base in a pocket within the enzyme.The mechanism of base flipping is not limited to DNA repair enzymes. In fact, it appears to be a generalized mechanism for catalytic DNA binding proteins (reviewed in Refs. 2 and 3). The first evidence that an enzyme flips a base extrahelical was revealed in the crystal structure of a DNA cytosine 5-methyltransferase, HhaI, complexed with its substrate DNA, in which the target cytosine was flipped out of the DNA helix and into the active site pocket of the enzyme where the methylation reaction can occur (4). A similar methyltransferase, HaeIII, has also been shown to move a base extrahelical (5), suggesting that this may be a conserved mechanism for this class of enzymes. As mentioned above, the two co...

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Role of the basic amino acid cluster and Glu-23 in pyrimidine dimer glycosylase activity of T4 endonuclease V.

Cited by 52 publications

References 15 publications

Intron Conservation in a UV-Specific DNA Repair Gene Encoded by Chlorella Viruses

Intron Conservation in a UV-Specific DNA Repair Gene Encoded by Chlorella Viruses

Temporal dynamics of Prochlorococcus ecotypes in the Atlantic and Pacific oceans

The Role of Base Flipping in Damage Recognition and Catalysis by T4 Endonuclease V

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