Dataset Information

Mismodeled purines: implicit alternates and hidden Hoogsteens.

ABSTRACT: Hoogsteen base pairs are seen in DNA crystal structures, but only rarely. This study tests whether Hoogsteens or other syn purines are either under-modeled or over-modeled, which are known problems for rare conformations. Candidate purines needing a syn/anti 180° flip were identified by diagnostic patterns of difference electron-density peaks. Manual inspection narrowed 105 flip candidates to 20 convincing cases, all at ?2.7?Å resolution. Rebuilding and refinement confirmed that 14 of these were authentic purine flips. Seven examples are modeled as Watson-Crick base pairs but should be Hoogsteens (commonest at duplex termini), and three had the opposite issue. Syn/anti flips were also needed for some single-stranded purines. Five of the 20 convincing cases arose from an unmodeled alternate duplex running in the opposite direction. These are in semi-palindromic DNA sequences bound by a homodimeric protein and show flipped-purine-like difference peaks at residues where the palindrome is imperfect. This study documents types of incorrect modeling which are worth avoiding. However, the primary conclusions are that such mistakes are infrequent, the bias towards fitting anti purines is very slight, and the occurrence rate of Hoogsteen base pairs in DNA crystal structures remains unchanged from earlier estimates at ?0.3%.

SUBMITTER: Hintze BJ

PROVIDER: S-EPMC5633910 | biostudies-literature | 2017 Oct

REPOSITORIES: biostudies-literature

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Mismodeled purines: implicit alternates and hidden Hoogsteens.

Hintze Bradley J BJ Richardson Jane S JS Richardson David C DC

Acta crystallographica. Section D, Structural biology 20171002 Pt 10

Hoogsteen base pairs are seen in DNA crystal structures, but only rarely. This study tests whether Hoogsteens or other syn purines are either under-modeled or over-modeled, which are known problems for rare conformations. Candidate purines needing a syn/anti 180° flip were identified by diagnostic patterns of difference electron-density peaks. Manual inspection narrowed 105 flip candidates to 20 convincing cases, all at ≤2.7 Å resolution. Rebuilding and refinement confirmed that 14 of these were ...[more]

PMID: 28994414

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Project description:Chlamydomonas reinhardtii is a model green microalga capable of heterotrophic growth on acetic acid but not fatty acids, despite containing a full complement of genes for β-oxidation. Recent reports indicate that the alga preferentially sequesters, rather than breaks down, lipid acyl chains as a means to rebuild its membranes rapidly. Here, we assemble a list of potential Chlamydomonas peroxins (PEXs) required for peroxisomal biogenesis to suggest that C. reinhardtii has a complete set of peroxisome biogenesis factors. To determine involvements of the peroxisomes in the metabolism of exogenously added fatty acids, we examined transgenic C. reinhardtii expressing fluorescent proteins fused to N- or C-terminal peptide of peroxisomal proteins, concomitantly with fluorescently labeled palmitic acid under different trophic conditions. We used confocal microscopy to track the populations of the peroxisomes in illuminated and dark conditions, with and without acetic acid as a carbon source. In the cells, four major populations of compartments were identified, containing: (1) a glyoxylate cycle enzyme marker and a protein containing peroxisomal targeting signal 1 (PTS1) tripeptide but lacking the fatty acid marker, (2) the fatty acid marker alone, (3) the glyoxylate cycle enzyme marker alone, and (4) the PTS1 marker alone. Less than 5% of the compartments contained both fatty acid and peroxisomal markers. Statistical analysis on optically sectioned images found that C. reinhardtii simultaneously carries diverse populations of the peroxisomes in the cell and modulates peroxisomal contents based on light conditions. On the other hand, the ratio of the compartment containing both fatty acid and peroxisomal markers did not change significantly regardless of the culture conditions. The result indicates that β-oxidation may be only a minor occurrence in the peroxisomal population in C. reinhardtii, which supports the idea that lipid biosynthesis and not β-oxidation is the primary metabolic preference of fatty acids in the alga.

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Mismodeled purines: implicit alternates and hidden Hoogsteens.

Publications

Mismodeled purines: implicit alternates and hidden Hoogsteens.

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