Characterization of the Conformational Binding of N-methyl Mesoporphyrin IX with DNA Model Telomeric G-Quadruplex Forming Sequences

DNA G-quadruplexes are alternative secondary structures formed from guanine rich DNA sequences. G-quadruplexes are found most abundantly at the ends of chromosomes called telomeres, where they can ultimately prevent further tumor growth and progression. Thus, quadruplex interactive agents, or ligands that can bind to and stabilize DNA G-quadruplex structures, are of high interest for their potential chemotherapeutic abilities. One quadruplex interactive agent is the porphyrin N-methyl mesoporphyrin IX (NMM), which has high binding selectivity for quadruplex structures. While the NMM-quadruplex complex in crystallized form has been greatly studied, this complex in solution form is not as well studied. Through circular dichroism (CD), fluorescence spectroscopy, and fluorescence lifetime studies, we were able to examine the structural rearrangements and folding of the quadruplex involved with NMM binding in solution, as well as the NMM binding site on the parallel quadruplex. This has allowed us to evaluate NMM as a conformational probe for quadruplex folding and formation and uncover information about folding pathways of telomeric DNA. For our studies, we have used a series of model human telomeric HT4 oligonucleotides, (TTAGGG)4, where the fluorophore, 6-methylisoxanthopterin (6-MI), is strategically introduced at specific locations along the model telomeric DNA sequence through replacement of key guanosine residues. CD spectroscopic studies show that, upon addition of NMM, a switch of the telomeric conformation, from the mixed-hybrid strand solution conformation promoted in 100 mM KCl, to a parallel strand arrangement, occurs. CD studies also allowed us to identify two groups of 6-MI substitutions that either ‘tolerate’ the conformational switch (Group 1), resulting in NMM binding, or ‘lock’ the conformation in the mixed-hybrid (Group 2), with little NMM interaction. Interestingly, Group 1 (G1-, G4, G7-, and G-10- and G11-6MI-qDNA) members exist predominantly in the syn-rotameric base orientation in the mixed-hybrid conformation. In contrast, guanine bases in locations that form members of Group 2 (G5-, G6-, G9- and G-12-6MI-qDNA) are in the more usual anti-rotameric form. Examination of fluorescence signals associated with NMM binding confirms that Group 1 guanine substitutions allow formation of the required parallel conformation, while Group 2 substitutions show little or weak associations of NMM with the “locked” mixed-hybrid conformation. Examination of 6-MI fluorescence signals with addition of NMM showed that all guanine 6-MI substitutions (Group 1 or 2), with addition of low [NMM], resulted in small enhancements of fluorescence emission intensity. Knowing that the fluorescence of 6-MI is significantly quenched because of base stacking interactions, relief of intensity quenching suggests that the mixed-hybrid participates in a conformational backbone rearrangement that results in a relief of base interactions. Addition of high [NMM] resulted in varying decreases of fluorescence quenching for Group 1 members. Determined Stern-Volmer Constant (Ks) values for these 6-MI quadruplex sequences that allow for the switch suggest that NMM likely has a distribution of guanine residues that contribute to the binding site, likely involving diagonal side loops on the parallel conformation, although 5’-end-stacking in the solution phase may not be ruled out. Fluorescence lifetime studies clearly provided evidence for a static ground state quenching process. Here the 6-MI and NMM must be in physical contact to form a non-fluorescent ground-state complex. In conclusion, studies described here provide more about the nature of the NMM-qDNA complex in solution. First, NMM binds to the parallel conformation. Second, structural rearrangements (strand relaxation/opening) prior to the main mixed hybrid to parallel conformation transition are evident. Third, formation of the parallel conformation requires strands 3 and 4 to effectively unfold from the mixed hybrid quadruplex, change direction and re-fold, now forming new stabilizing interactions, including Hoogsteen base pairing. The parallel form of the quadruplex appears to provide a more condensed, crystal-like structure for NMM binding

Comparison of perceptions of the Canossian school system in three Canossian schools

This study aimed to identify and compare the perceptions of the administrators and teachers on the extent to which the Canossian School System is reflected in the Canossian schools in Laguna, school year 2009-2010.Using a researcher- developed instrumen

The photophysics and photochemistry of carotenoids {\it in vitro\/} and in light-harvesting pigment-protein complexes

Carotenoids supplement the light-capturing ability of (bacterio)chlorophyll in antenna pigment-protein complexes by harvesting light in the $\sim$425-500 nm visible region. Energy is then transferred from the carotenoids-to-(bacterio)chlorophyll via singlet energy transfer. To understand the molecular details of how carotenoids carry out their light-harvesting role, the spectroscopic properties of spheroidene and a series of spheroidene analogs with extents of $\pi$-electron conjugation ranging from 7 to 13 carbon-carbon double bonds were studied using steady-state and time resolved optical spectroscopy. Studies were conducted with the carotenoids in solution and incorporated into the B850 light-harvesting complex from Rhodobacter sphaeroides R-26.1. The spheroidene analogs used were 5\sp\prime,6\sp\prime-dihydro-7\sp \prime,8\sp\prime-didehydrospheroidene, 7\sp\prime,8\sp\prime-didehydrospheroidene, and 1\sp\prime,2\sp\prime-dihydro-3\sp\prime,4\sp \prime,7\sp\prime,8\sp\prime-tetradehydrospheroidene. These data, taken together with results from 3,4,7,8-tetrahydrospheroidene, 3,4,5,6-tetrahydrospheroidene, 3,4-dihydrospheroidene and spheroidene already published (DeCoster, B., et al. (1992) Biochim. Biophys. Acta 1102, 107-114.: Frank, H. A., et al. (1993) Chem. Phys. Lett. 207, 88-92.: Frank, H. A., et al. (1993) Photochem. Photobiol. 57, 49-55.: and Farhoosh, R., et al. (1994) Photosyn. Res. 42, 157-166.) provide a systematic series of molecules for understanding the molecular features that control energy transfer to bacteriochlorophyll in photosynthetic bacterial light-harvesting complexes. Based on the results of the in vitro studies it was hypothesized that transfer from the S\sb1 state of the carotenoid is more significant for the shorter molecules ($\le$10 carbon-carbon double bonds) whereas for longer carotenoids ($\u3e$10 carbon-carbon double bonds), transfer from the S\sb2 state is more important. After the behavior of the carotenoids in solution was established the carotenoids were incorporated into the B850 light-harvesting complex. Data obtained support the hypothesis that only carotenoids having 10 or less carbon-carbon double bonds transfer energy via their S\sb1 \rm(2\sp1A\sb{g}) states to bacteriochlorophyll to any significant degree. Energy transfer via the S\sb2 \rm(1\sp1B\sb{u}) state of the carotenoid becomes more important than the S\sb1 route as the number of conjugated carbon-carbon double bonds increase. The results also suggest that the S\sb2 state associated with the Q\rm\sb{x} transition of the B850 bacteriochlorophyll, is the most likely acceptor state for energy transfer originating from both the \rm2\sp1A\sb{g} and \rm 1\sp1B\sb{u} states of all carotenoids.

Some Ectoparasites from Philippine Owls (Strigiformes: Strigidae) with Description of a New Louse Species, Kurodaia (Conciella) theresamunditae Desamero & Eduardo (Amblycera: Menoponidae) *FOR CORRESPONDENCE

ABSTRACT Two species of Philippine owls, Otus megalotis Walden, 1875 (Philippine scops-owl) and Bubo philippensis Kaup, 1851 (Philippine eagle-owl) (Strigidae), kept at the Rescue Center of the Protected Areas and Wildlife Bureau (PAWB), Quezon City, Philippines were examined for ectoparasites during the period June 2009-February 2010. Lice and mites were found on the two species of owls. Identification of the lice specimens revealed four species, three previously known and one new to science. These were Strigiphilus heterogenitalis from Otus megalotis, Colpocephalum turbinatum from both Otus megalotis and Bubo philippensis and Strigiphilus ketupae from the latter owl species. Kurodaia (Conciella) theresamunditae Desamero & Eduardo is described and illustrated as a new species from Otus megalotis. It is characterized by having male genitalia and the associated genital sclerite different from those of the other species of the genus and subgenus to which it belongs. The mite specimens consisted of a single species identified as Dermonoton longiventer. The occurrence of S. heterogenitalis and H. ketupae on Otus megalotis and Bubo philippensis, respectively constitutes new host records for the species in the Philippines. The presence of Colpocephalum turbinatum on both owl species in this study extends the host range of the species. Dermonoton longiventer is reported for the first time in the Philippines

Large Scale Dynamics of the Michaelis Complex in Bacillus stearothermophilus Lactate Dehydrogenase Revealed by a Single-Tryptophan Mutant Study

 An information technology is suggested to provide the automatic development of simulated models by converting graphic models of business process, got by other programs for modeling. Got simulated models can be used for the synthetic environment construction of information security systems to define more exactly models features and characteristics. Developed information technology can be as base for distributed modeling systems constructing within the limits of realization HLA. Рассмотрена технология формализированного представления имитационных моделей систем информационной безопасности на основе конвертации формализированных описаний моделей, полученных с помощью других программ моделирования бизнес-процессов синтетического окружения систем в XML-файлы. Имитационные модели могут использоваться для построения синтетического окружения систем информационной безопасности для уточнения их свойств и характеристик. Разработанная информационная технология может служить основой для построения распределенного моделирования в рамках реализации HLA. Запропоновано технологію до формалізованого подання імітаційних моделей систем інформаційної безпеки на основі конвертації формалізованих описів моделей, отриманих за допомогою інших програм моделювання бізнес-процесів синтетичного оточення систем в XML-файли

Insights on the Functional Role of Beta-Glucans in Fungal Immunity Using Receptor-Deficient Mouse Models

Understanding the host anti-fungal immunity induced by beta-glucan has been one of the most challenging conundrums in the field of biomedical research. During the last couple of decades, insights on the role of beta-glucan in fungal disease progression, susceptibility, and resistance have been greatly augmented through the utility of various beta-glucan cognate receptor-deficient mouse models. Analysis of dectin-1 knockout mice has clarified the downstream signaling pathways and adaptive effector responses triggered by beta-glucan in anti-fungal immunity. On the other hand, assessment of CR3-deficient mice has elucidated the compelling action of beta-glucans in neutrophil-mediated fungal clearance, and the investigation of EphA2-deficient mice has highlighted its novel involvement in host sensing and defense to oral mucosal fungal infection. Based on these accounts, this review focuses on the recent discoveries made by these gene-targeted mice in beta-glucan research with particular emphasis on the multifaceted aspects of fungal immunity