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001-es BibID:	BIBFORM023456
Első szerző:	Vámosi György (biofizikus)
Cím:	Fluorescence characteristics of 5-carboxytetramethylrhodamine linked covalently to the 5' end of oligonucleotides : multiple conformers of single-stranded and double-stranded dye-DNA complexes / György Vámosi, Christoph Gohlke, Robert M. Clegg
Dátum:	1996
ISSN:	0006-3495
Megjegyzések:	Fluorescence steady-state and lifetime experiments have been carried out on duplex and single-stranded DNA molecules labeled at the 5' ends with 5-carboxytetramethylrhodamine (TMRh). The temperature and ionic strength of the solutions were varied over large ranges. The results reveal at least three well-defined states of the TMRh-DNA molecules for the single-stranded as well as for the double-stranded DNA molecules. Two states are fluorescent, with lifetimes in the range of 0.5-1 ns and 2.5-3 ns. A third state of TMRh-DNA does not fluoresce (a dark species of TMRh-DNA). The distribution of the TMRh-DNA molecules among these three states is strongly temperature and ionic strength dependent. Estimates are made of some reaction parameters of the multistate model. The results are discussed in terms of the photophysics of TMRh, and consequences of the multiple conformers of TMRh-DNA for studies involving fluorescence studies with TMRh-labeled DNA are considered.
Tárgyszavak:	Természettudományok Biológiai tudományok idegen nyelvű folyóiratközlemény külföldi lapban
Megjelenés:	Biophysical Journal. - 71 : 2 (1996), p. 972-994. -
További szerzők:	Gohlke, Christoph Clegg, Robert M.
Internet cím:	Szerző által megadott URL Intézményi repozitóriumban (DEA) tárolt változat
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001-es BibID:	BIBFORM023462
Első szerző:	Vámosi György (biofizikus)
Cím:	Helix-coil transition of a four-way DNA junction observed by multiple fluorescence parameters / Vámosi G., Clegg R. M.
Dátum:	2008
ISSN:	1520-6106
Megjegyzések:	The thermal denaturation of immobile four-way DNA ("Holliday-") junctions with 17 base pair arms was studied via fluorescence spectroscopic measurements. Two arms of the molecule were labeled at the 5'-end with fluorescein and tetramethylrhodamine, respectively. Melting was monitored by the fluorescence intensity of the dyes, the fluorescence anisotropy of tetramethylrhodamine, and Forster resonance energy transfer (FRET) between fluorescein and rhodamine. To fit the thermal denaturation curves of the four-way junctions, two basic thermodynamic models were tested: (1) all-or-none transitions assuming a molecularity of one, two, or four and (2) a statistical "zipper" model. The all-or-none models correspond to reaction mechanisms assuming that the cooperative melting unit (that is, the structure changing from complete helix to complete coil) consists of (1) one arm, (2) two neighboring arms (which have one continuous strand common to the two arms), or (3) all four arms. In each case, the melting of the cooperative unit takes place in a single step. The tetramolecular reaction model (four-arm melting) yielded unrealistically low van't Hoff enthalpy and entropy values, whereas the monomolecular model (one-arm melting) resulted in a poor fit to the experimental data. The all-or-none bimolecular (two neighboring arm model) fit gave intermediate standard enthalpy change (Delta H) values between those expected for the melting of a duplex with a total length between the helix lengths of one and two arms (17 and 34 base pairs). Simulations according to the zipper model fit the experimental curves best when the length of the simulated duplex was assumed to be 34 base pairs, the length of a single strand. This suggests that the most important parameter determining the melting behavior of the molecule is the end-to-end distance of the strands (34 bases) rather than the length of the individual arms (17 base pairs) and that the equilibrium concentration of partially denatured intermediate states has to be taken into account. These findings are in good agreement with results obtained for three-way DNA junctions (Stuhmeier, F.; Lilley, D. M.; Clegg, R. M. Biochemistry 1997, 36, 13539). An interesting result is that the extent-of-melting curves derived from the fluorescence intensity and anisotropy nearly agree, whereas the curve derived from the FRET data shows a change prior to the melting. This may be an indication of a conformational change leaving the doublestranded structure intact but changing the end-to-end distance of the different arms in a way consistent with the transition to the extended square configuration (Clegg, R. M.; Murchie, A. I.; Lilley, D. M. Biophys. J. 1994, 66, 99) of this branched molecule.
Tárgyszavak:	Természettudományok Biológiai tudományok idegen nyelvű folyóiratközlemény külföldi lapban DNA melting
Megjelenés:	Journal Of Physical Chemistry B. - 112 : 41 (2008), p. 13136-13148. -
További szerzők:	Clegg, Robert M.
Pályázati támogatás:	T48745 OTKA
Internet cím:	Szerző által megadott URL DOI Intézményi repozitóriumban (DEA) tárolt változat
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001-es BibID:	BIBFORM004641
Első szerző:	Vámosi György (biofizikus)
Cím:	The helix-coil transition of DNA duplexes and hairpins observed by multiple fluorescence parameters / Vamosi, G., Clegg, R. M.
Dátum:	1998
ISSN:	006-2960 (Print)
Megjegyzések:	The thermal denaturation of 8-20-bp DNA duplexes labeled with fluorescein and tetramethylrhodamine at opposing 5'-ends was investigated by monitoring the fluorescence intensity of the dyes, the fluorescence anisotropy of tetramethylrhodamine, the fluorescence resonance energy transfer between fluorescein and rhodamine, and, for the 20-bp duplex, the UV absorption. Melting experiments with the single strands of the duplexes revealed that the single strands can form hairpins stabilized by only a few base pairs. The thermal denaturation curves of the duplexes were fitted well to an extended all-or-none model assuming that only the fully base-paired duplex, the maximally base-paired hairpin, and the random coil conformations are present simultaneously. The extent-of-melting versus temperature curves derived from the different spectroscopic parameters are nearly identical, provided that the analysis of the baselines is carried out correctly; the DeltaH and DeltaS of the dissociation compare well with predictions based on nearest neighbor interaction values available in the literature. Our results imply that for all the oligonucleotides other than the 34-bp oligomer, no partially melted intermediates other than hairpins are present in the reaction mixture in amounts that can be detected by our methods. The melting of the hairpins was also studied directly using single-stranded oligonucleotides. The melting of a 34-bp duplex can be accounted for by a statistical zipper model
Tárgyszavak:	Orvostudományok Elméleti orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban analysis chemical synthesis chemistry Dna Single-Stranded Dyes Energy Transfer Entropy Fluorescein Fluorescence Fluorescence Polarization Fluorescence Resonance Energy Transfer Fluorescent Dyes Heat methods Models,Chemical Nucleic Acid Conformation Nucleic Acid Denaturation Nucleic Acid Heteroduplexes Oligonucleotides Research Rhodamines Spectrometry Support Temperature Thermodynamics
Megjelenés:	Biochemistry. - 37 : 40 (1998), p. 14300-14316. -
További szerzők:	Clegg, Robert M.
Internet cím:	elektronikus változat DOI
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