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81.
Marilynn A. Larson Mark A. Griep Rafael Bressani Kiran Chintakayala Panos Soultanas Steven H. Hinrichs 《Nucleic acids research》2010,38(20):7167-7178
Bacterial primase is stimulated by replicative helicase to produce RNA primers that are essential for DNA replication. To identify mechanisms regulating primase activity, we characterized primase initiation specificity and interactions with the replicative helicase for gram-positive Firmicutes (Staphylococcus, Bacillus and Geobacillus) and gram-negative Proteobacteria (Escherichia, Yersinia and Pseudomonas). Contributions of the primase zinc-binding domain, RNA polymerase domain and helicase-binding domain on de novo primer synthesis were determined using mutated, truncated, chimeric and wild-type primases. Key residues in the β4 strand of the primase zinc-binding domain defined class-associated trinucleotide recognition and substitution of these amino acids transferred specificity across classes. A change in template recognition provided functional evidence for interaction in trans between the zinc-binding domain and RNA polymerase domain of two separate primases. Helicase binding to the primase C-terminal helicase-binding domain modulated RNA primer length in a species-specific manner and productive interactions paralleled genetic relatedness. Results demonstrated that primase template specificity is conserved within a bacterial class, whereas the primase–helicase interaction has co-evolved within each species. 相似文献
82.
83.
Construction and characterization of mismatch-containing circular DNA molecules competent for assessment of nick-directed human mismatch repair in vitro 下载免费PDF全文
The ability of cell-free extracts to correct DNA mismatches has been demonstrated in both prokaryotes and eukaryotes. Such an assay requires a template containing both a mismatch and a strand discrimination signal, and the multi-step construction process can be technically difficult. We have developed a three-step procedure for preparing DNA heteroduplexes containing a site-specific nick. The mismatch composition, sequence context, distance to the strand signal, and the means for assessing repair in each strand are adjustable features built into a synthetic oligonucleotide. Controlled ligation events involving three of the four DNA strands incorporate the oligonucleotide into a circular template and generate the repair-directing nick. Mismatch correction in either strand of a prototype G·T mismatch was achieved by placing a nick 10–40 bp away from the targeted base. This proximity of nick and mismatch represents a setting where repair has not been well characterized, but the presence of a nick was shown to be essential, as was the MSH2/MSH6 heterodimer, although low levels of repair occurred in extract defective in each protein. All repair events were inhibited by a peptide that interacts with proliferating cell nuclear antigen and inhibits both mismatch repair and long-patch replication. 相似文献
84.
DM Woolley 《The Journal of cell biology》1977,75(3):851-865
Motile spermatozoa from the golden hamster have been arrested by rapid freezing and then fixed with glutaraldehyde at low temperature after substitution with ethylene glycol. As far as can be judged, the flagellar waveforms thus stabilized are similar to those seen in living sperm; in contrast, fixation in glutaraldehyde, without prior freezing, induces agonal changes in flagellar conformation. The characteristics waveform after freeze substitution contains three bends. Approx. half of these flagella are entirely planar. The rest are three dimensional, with the third bend displaced in a regular way from the plane containing the proximal two bends. From the geometry of these flagella, it is concluded that the plane of action of a given bending cycle undergoes a clockwise twist (from a forward viewpoint) as the cycle is succeeded by new bending cycles. This "twisted plane" undulation is quite different from helical movement. The twisting seems to occur abruptly, between cycles, as if each bending cycle has a preferred plane of action. The mechanism underlying the twisting is uncertain. However, on the basis of the angular displacements between the preferred planes, and the findings from electron microscopy, the following idea is presented as a working hypothesis: that, if the most proximal plane of bending is topographically determined by peripheral doublet 1, then successive distal planes of action are influenced predominantly by doublets 2, 3, etc., in clockwise sequence. The merits and weaknesses of this hypothesis are discussed. 相似文献
85.
86.
Codon usage patterns in Nematoda: analysis based on over 25 million codons in thirty-two species 总被引:1,自引:0,他引:1
Makedonka Mitreva Michael C Wendl John Martin Todd Wylie Yong Yin Allan Larson John Parkinson Robert H Waterston James P McCarter 《Genome biology》2006,7(8):R75-19
Background
Codon usage has direct utility in molecular characterization of species and is also a marker for molecular evolution. To understand codon usage within the diverse phylum Nematoda, we analyzed a total of 265,494 expressed sequence tags (ESTs) from 30 nematode species. The full genomes of Caenorhabditis elegans and C. briggsae were also examined. A total of 25,871,325 codons were analyzed and a comprehensive codon usage table for all species was generated. This is the first codon usage table available for 24 of these organisms. 相似文献87.
88.
The ITS sequences of Acropora spp. are the shortest so far identified in
any metazoan and are among the shortest seen in eukaryotes; ITS1 was 70-80
bases, and ITS2 was 100-112 bases. The ITS sequences were also highly
variable, but base composition and secondary structure prediction indicate
that divergent sequence variants are unlikely to be pseudogenes. The
pattern of variation was unusual in several other respects: (1) two
distinct ITS2 types were detected in both A. hyacinthus and A. cytherea,
species known to hybridize in vitro with high success rates, and a putative
intermediate ITS2 form was also detected in A. cytherea; (2) A. valida was
found to contain highly (29%) diverged ITS1 variants; and (3) A.
longicyathus contained two distinct 5.8S rDNA types. These data are
consistent with a reticulate evolutionary history for the genus Acropora.
相似文献
89.
Atomistic molecular dynamics simulations and structural bioinformatics tools enable the identification of the exact mode of interaction between model pulmonary surfactant components. Two nanosecond long simulations of the N-terminal region of human surfactant protein-B (SP-B(1-25)) in dipalmitoylphosphatidylcholine (DPPC) and dipalmitoylphosphatidylglycerol (DPPG) monolayers of different lipid surface densities reveal the preferential affinity of SP-B(1-25) for anionic phospholipids. In particular, arginine 12 and lysine 24 interact strongly and with high specificity with the phosphate group of the DPPG lipids, stabilizing the position, the orientation, and the secondary structure of the peptide in the monolayer. The peptide lies at an oblique angle to the interfacial plane, ranging between 47 degrees and 62 degrees, increasing with decreasing lipid surface density. In DPPC monolayers the interaction is largely determined by hydrophobic interactions. The non-specific nature of DPPC-SP-B(1-25) interactions allows for significant flexibility in the topology of the peptide in the lipid matrix. Bioinformatics tools are employed to generalize the simulation results to the sequences of SP-B(1-25) in other organisms. The importance of specific residues, and the role of the largely helical and amphiphilic nature of the peptide in the functionality of SP-B(1-25) are established. The synergy of classical mechanics tools with bioinformatics methods greatly enhances the molecular-level interpretation of pulmonary surfactant action and facilitates the development of design rules for synthetic surfactant analogues. 相似文献
90.
VLJ Whitehall TD Dumenil DM McKeone CE Bond ML Bettington RL Buttenshaw L Bowdler GW Montgomery LF Wockner BA Leggett 《Epigenetics》2014,9(11):1454-1460
The CpG Island Methylator Phenotype (CIMP) is fundamental to an important subset of colorectal cancer; however, its cause is unknown. CIMP is associated with microsatellite instability but is also found in BRAF mutant microsatellite stable cancers that are associated with poor prognosis. The isocitrate dehydrogenase 1 (IDH1) gene causes CIMP in glioma due to an activating mutation that produces the 2-hydroxyglutarate oncometabolite. We therefore examined IDH1 alteration as a potential cause of CIMP in colorectal cancer. The IDH1 mutational hotspot was screened in 86 CIMP-positive and 80 CIMP-negative cancers. The entire coding sequence was examined in 81 CIMP-positive colorectal cancers. Forty-seven cancers varying by CIMP-status and IDH1 mutation status were examined using Illumina 450K DNA methylation microarrays. The R132C IDH1 mutation was detected in 4/166 cancers. All IDH1 mutations were in CIMP cancers that were BRAF mutant and microsatellite stable (4/45, 8.9%). Unsupervised hierarchical cluster analysis identified an IDH1 mutation-like methylation signature in approximately half of the CIMP-positive cancers. IDH1 mutation appears to cause CIMP in a small proportion of BRAF mutant, microsatellite stable colorectal cancers. This study provides a precedent that a single gene mutation may cause CIMP in colorectal cancer, and that this will be associated with a specific epigenetic signature and clinicopathological features. 相似文献