Abstract:
:Targeting damage to specific sites in the genome represents an attractive approach to manipulating gene function in mammalian cells. To test the applicability of triple-helix formation as a means for achieving precisely timed site-specific damage within a mammalian gene, a triplex-forming oligodeoxyribonucleotide (TFO) that binds with high affinity to a specific site within the hamster adenine phosphoribosyltransferase (APRT) gene was modified with the photochemically reactive psoralen derivative 4'-(hydroxymethyl)-4,5',8-trimethylpsoralen (HMT). The modified TFO, psorTFO1, bound with high affinity to a target site within intron 1 of the APRT gene. Upon irradiation, photomonoadducts (i.e., covalent adducts of psorTFO1 to one strand of the target duplex) were formed with high efficiency (approximately 50%). Introduction of 5'-TpA sequences (the preferred site for psoralen-induced photo-cross-links) at or near the triplex junction leads to increased efficiency of total photoadduct formation and to efficient formation of products that had the electrophoretic mobility on denaturing PAGE expected for three-stranded photo-cross-links (i.e., products containing psorTFO1 covalently linked to both strands of the duplex). Their identities as cross-links were verified by (1) identical electrophoretic mobility of products formed with either duplex strand radiolabeled and (2) coprecipitation of the radiolabeled duplex strand with its complementary biotinylated strand following denaturation. In addition, the cross-links were completely reversible upon irradiation at 254 nm, as expected for psoralen-mediated cross-links. The yield and distribution of photoadducts depended on 5'-TpA position. The most efficient photoadduct formation (approximately 90%) and photo-cross-link formation (approximately 90% of total photoadducts) were observed for a 5'-TpA adjacent to the triplex junction, with significant, but lower, cross-linking efficiency within three base pairs of the junction. Molecular models of the psoralen-conjugated triplex with its six-carbon linker suggested a simple explanation for this distance dependence: facile intercalation near the triplex/duplex junction, with increasing strain required for intercalation at more distant sites. These results indicate that psorTFO1 allows for DNA damage with high precision and high efficiency, and the likely proportion of monoadducts and cross-links can be estimated from the target sequence.
journal_name
Biochemistryjournal_title
Biochemistryauthors
Vasquez KM,Wensel TG,Hogan ME,Wilson JHdoi
10.1021/bi960881fsubject
Has Abstractpub_date
1996-08-20 00:00:00pages
10712-9issue
33eissn
0006-2960issn
1520-4995pii
bi960881fjournal_volume
35pub_type
杂志文章相关文献
BIOCHEMISTRY文献大全abstract::The solution structure of Cucurbita maxima trypsin inhibitor-V (CMTI-V), which is also a specific inhibitor of the blood coagulation protein, factor XIIa, was determined by 1H NMR spectroscopy in combination with a distance-geometry and simulated annealing algorithm. Sequence-specific resonance assignments were made f...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00015a034
更新日期:1995-04-18 00:00:00
abstract::The bacterium Caulobacter crescentus is known to attach irreversibly to underwater surfaces by utilizing an adhesive structure called the holdfast, which exhibits the greatest known adhesive strength of any organism. The very small size of the holdfast (∼400 nm wide and ∼40 nm high) has made direct chemical analysis d...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/acs.biochem.0c00625
更新日期:2020-09-22 00:00:00
abstract::Decarboxylated S-adenosylmethionine was found to be a substrate for the nuclear acetyltransferases that act on polyamines and on histones. The rate of acetylation of decarboxylated S-adenosylmethionine was more than twice that of spermidine at saturating substrate concentrations, and decarboxylated S-adenosylmethionin...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00350a016
更新日期:1986-01-28 00:00:00
abstract::In Neurospora crassa, the expression of the nit-3 gene (nitrate reductase) is dependent upon nitrogen derepression and nitrate induction and is regulated by two positive-acting transcription factors, NIT2 and NIT4, and a negative regulator, NMR. The presence of a tightly linked cluster of NIT2 and NIT4 binding sites s...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi980224i
更新日期:1998-08-04 00:00:00
abstract::Time-resolved resonance Raman (TR(3)) and time-resolved step-scan (TRS(2)) FTIR spectroscopies have been used to probe the structural dynamics at the heme b(3) proximal and distal sites after carbon monoxide photolysis from fully reduced CO-bound nitric oxide reductase. The Raman spectra of the transient species exhib...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi035289m
更新日期:2003-12-23 00:00:00
abstract::The present paper reports a study on the equilibria and kinetics of the acid-alkaline transition and the azide binding reaction by ferric Aplysia myoglobin. A single completely reversible spectrophotometric titration curve is found over the pH range from similar to 5 to similar to 9, with an apparent pK equals to 7.5 ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00679a006
更新日期:1975-04-22 00:00:00
abstract::Monomeric sarcosine oxidase (MSOX) is a prototypical member of a recently recognized family of amine-oxidizing enzymes that all contain covalently bound flavin. Mutation of the covalent flavin attachment site in MSOX produces a catalytically inactive apoprotein (apoCys315Ala) that forms an unstable complex with FAD (K...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi0607352
更新日期:2006-08-08 00:00:00
abstract::An antibody population that reacted with the high-affinity lysine binding site of human plasminogen was elicited by immunizing rabbits with an elastase degradation product containing kringles 1-3 (EDP I). This antibody was immunopurified by affinity chromatography on plasminogen-Sepharose and elution with 0.2 M 6-amin...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00370a028
更新日期:1986-11-04 00:00:00
abstract::The biochemical characteristics of the formation and disappearance of intercalator-induced DNA double-strand breaks (DSB) were studied in nuclei from mouse leukemia L1210 cells by using filter elution methodology [Bradley, M. O., & Kohn, K.W. (1979) Nucleic Acids Res. 7, 793-804]. The three intercalators used were 4'-...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00309a013
更新日期:1984-07-03 00:00:00
abstract::The three-dimensional structure of the fatty-acid-binding protein isolated from the flight muscle of the desert locust Schistocerca gregaria has been solved and refined to a crystallographic R-value of 18.5% for all measured X-ray data from 30.0- to 2.2-A resolution. Crystals employed in the investigation were grown f...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00207a004
更新日期:1994-10-18 00:00:00
abstract::In the anaerobic sulfate-reducing bacterium Desulfovibrio vulgaris Hildenborough (DvH), the genome sequencing revealed the presence of three operons encoding formate dehydrogenases. fdh1 encodes an alphabetagamma trimeric enzyme containing 11 heme binding sites; fdh2 corresponds to an alphabetagamma trimeric enzyme wi...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi0515366
更新日期:2005-11-15 00:00:00
abstract::The enzyme hydrogenase, from the photosynthetic bacterium Chromatium, was purified to homogeneity after solubilization of the particulate enzyme with deoxycholate. The purification procedure included ammonium sulfate fractionation, treatment with manganous phosphate gel, heating at 63 degrees, DEAE-cellulose chromatog...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00683a001
更新日期:1975-06-17 00:00:00
abstract::We have studied the physical and functional linkages of heterologously expressed human 5-HT1A receptors to G protein alpha-subunits in HeLa and CHO-K1 cells. HeLa cells expressed immunoreactivity to G(i) proteins with an apparent rank order of G(i) alpha 3 (approximately 1 pmol/mg of protein) > G(i) alpha 1 (approxima...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00092a016
更新日期:1993-10-19 00:00:00
abstract::The effects of structural modifications of the flavin mononucleotide (FMN) prosthetic group of Clostridium pasteurianum flavodoxin on the kinetics of electron transfer to the oxidized form (from 5-deazariboflavin semiquinone produced by laser flash photolysis) and from the semiquinone form (to horse heart cytochrome c...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00281a034
更新日期:1983-06-07 00:00:00
abstract::pH-dependent Ca2+ and Na+ transport via Na-Ca+K exchange was measured in isolated and purified bovine retinal rod outer segments (ROS). Ca2+ transport was measured (1) as 45Ca uptake via reverse Na-Ca+K exchange or via Ca-Ca exchange and (2) as a Ca(2+)-induced rise in cytosolic free Ca2+ concentration via reverse Na-...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00021a045
更新日期:1995-05-30 00:00:00
abstract::Murine epidermal growth factor (EGF), a 53 amino acid protein, has been modified by enzymic digestion, site-specific chemical reactions, and recombinant DNA technology. After trypsin digestion the EGF derivatives EGF1-48 (called EGF-T) and EGF1-45 (called EGF-T2) were separated from the residual EGF and the C-terminal...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00414a005
更新日期:1988-07-12 00:00:00
abstract::The technique of fluorescence recovery after photobleaching is used to perform an extensive study of the lateral diffusion of a phospholipid probe in the binary mixture dimyristoylphosphatidylcholine/cholesterol, above the melting temperature of the phospholipid. In the regions of the phase diagram where a single liqu...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00144a013
更新日期:1992-07-28 00:00:00
abstract::We have recently described a procedure for the purification of microtubule associated protein 1B (MAP1B) from calf brain [Pedrotti, B., & Islam K. (1995) Cell Motil. Cytoskeleton 30, 301-309], and this study further characterizes the purified protein and its interaction with microtubules. We show that purified MAP1B (...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi951314f
更新日期:1996-03-05 00:00:00
abstract::Resonance Raman spectra of the photosynthetic reaction center isolated from the green bacterium Chloroflexus aurantiacus have been obtained with excitation in the near-infrared absorption bands of the special pair (P) and the accessory bacteriochlorophyll (B) using shifted-excitation Raman difference spectroscopy (SER...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00015a044
更新日期:1995-04-18 00:00:00
abstract::Only beta-beta cross-links form when the alpha(3)(betaE(395)C)(3)gammaK(36)C (MF(1) residue numbers) double mutant subcomplex of TF(1), the F(1)-ATPase from the thermophilic Bacillus PS3, is slowly inactivated with CuCl(2) in the presence or absence of MgATP. The same slow rate of inactivation and extent of beta-beta ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi036058i
更新日期:2004-03-09 00:00:00
abstract::Through a ligand-directed tosyl (LDT) chemistry strategy using the synthetic acetogenin ligand AL1, we succeeded in the pinpoint alkynylation (-C≡CH) of Asp160 in the 49 kDa subunit of bovine complex I, which may be located in the inner part of the putative quinone binding cavity of the enzyme [Masuya, T., et al. (201...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi501342w
更新日期:2014-12-16 00:00:00
abstract::We measured the time course of skeletal muscle actin polymerization at different actin concentrations. In 0.1 M KCl with 1 mM Mg2+, log/log plots of the rate of the early, slow phase of polymerization vs. actin concentration were linear with slopes from 1.0 to 1.3. Computer-assisted calculations of similar curves from...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00278a021
更新日期:1983-04-26 00:00:00
abstract::The periplasmic protein CusF, as a part of the CusCFBA efflux complex, plays a role in resistance to elevated levels of copper and silver in Escherichia coli. Although homologues have been identified in other Gram-negative bacteria, the substrate of CusF and its precise role in metal resistance have not been described...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi0612622
更新日期:2006-09-19 00:00:00
abstract::Mouse photoreceptor function and survival critically depend on Ca(2+)-regulated retinal membrane guanylyl cyclase (RetGC), comprised of two isozymes, RetGC1 and RetGC2. We characterized the content, catalytic constants, and regulation of native RetGC1 and RetGC2 isozymes using mice lacking guanylyl cyclase activating ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi200491b
更新日期:2011-06-28 00:00:00
abstract::The binding of 8-anilino-1-naphthalenesulfonate (ANS) to ciliary dynein ATPase leads to a marked increase in the dye's fluorescence intensity, accompanied by a blue shift in the observed fluorescence emission maximum. We found that dynein has 37 +/- 3 ANS binding sites and that experimentally applied ANS concentration...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00347a012
更新日期:1985-12-17 00:00:00
abstract::The cyanobacterium Synechocystis sp. PCC 6803 (S6803) expresses a two-on-two globin in which His46 (distal side) and His70 (proximal) function as heme iron axial ligands. His46 can be displaced by O2, CO, and CN-, among others, whereas His70 is not labile under native conditions. The residue preceding the proximal his...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi060691x
更新日期:2006-09-26 00:00:00
abstract::A new series of peptide inhibitors of human Factor VIIa (FVIIa) has been identified and affinity matured from naive and partially randomized peptide phage libraries selected against the immobilized tissue factor x Factor VIIa (TF x FVIIa) complex. These "A-series" peptides contain a single disulfide bond and a 13-resi...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi010591l
更新日期:2001-08-14 00:00:00
abstract::The NADH absorbance spectrum of nicotinoprotein (NADH-containing) alcohol dehydrogenase from Amycolatopsis methanolica has a maximum at 326 nm. Reduced enzyme-bound pyridine dinucleotide could be reversibly oxidized by acetaldehyde. The fluorescence excitation spectrum for NADH bound to the enzyme has a maximum at 325...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi972115u
更新日期:1998-03-03 00:00:00
abstract::The Wilson disease copper-transporting ATPase plays a critical role in the intracellular trafficking of copper. Mutations in this protein lead to the accumulation of a toxic level of copper in the liver, kidney, and brain followed by extensive tissue damage and death. The ATPase has a novel amino-terminal domain ( app...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi992222j
更新日期:2000-02-22 00:00:00
abstract::Coiled coils are simple models for studying the association of two polypeptide chains to form a folded protein. Previous work has shown that the folding of a coiled coil can be described by a two-state transition between two unfolded monomeric peptide chains and a folded coiled coil dimer. Here we report the thermodyn...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi002161l
更新日期:2001-03-27 00:00:00