共查询到20条相似文献,搜索用时 31 毫秒
1.
D V Pyshnyi I Pyshnaya A Levina E Goldberg V Zarytova D Knorre E Ivanova 《Journal of biomolecular structure & dynamics》2001,19(3):555-570
Contiguous stacking hybridization of oligodeoxyribonucleotides with DNA as template was investigated using three types of complexes: oligonucleotide contiguously stacked with the stem of the preformed minihairpin (complexes I), oligonucleotide tandems containing two (complexes II) or three (complexes III) short oligomers with a common DNA template. Enthalpy Delta H degrees and entropy Delta S degrees of the coaxial stacking of adjacent duplexes were determined for GC/G*pC, GT/A*pC, AC/G*pT, AT/A*pT, CT/A*pG, AG/C*pT, AA/T*pT and TT/A*pA nicked (*) dinucleotide base pairs. The maximal efficiency of co-operative interaction was found for the GC/G*pC interface (Delta G degrees(NN/N*pN)=-2.7 kcal/mol) and the minimal one for the AA/T*pT interface (Delta G degrees(NN/N*pN)=-1.2 kcal/mol) at 37 degrees C. As a whole, the efficiency of the base pairs interaction Delta G degrees(NN/N*pN) in the nick is not lower than that within the intact DNA helix (Delta G degrees(NN/NN)).These observed Delta G degrees(NN/N*pN) values are proposed may include the effect of the partial removal of fraying at the adjacent helix ends additionally to the effect of the direct stacking of the terminal base pairs in the duplex junction (Delta G degrees(NN/NN). The thermodynamic parameters have been found to describe adequately the formation of all tandem complexes of the II and III types with oligonucleotides of various length and hybridization properties. The performed thermodynamic analysis reveals features of stacking oligonucleotide hybridization which allow one to predict the temperature dependence of association of oligonucleotides and the DNA template within tandem complexes as well as to determine optimal concentration for formation of these complexes characterized by high co-operativity level. 相似文献
2.
《Nucleosides, nucleotides & nucleic acids》2013,32(6-7):1057-1064
Contiguous stacking hybridization of oligodeoxyribonucleotides with a stem of preformed minihairpin structure of a DNA template was studied with the use of UV‐melting technique. It was shown that the free‐energy of the coaxial stacking interaction (ΔG°ST at 37°C, 1 M NaCl, pH 7.4) at the complementary interface XA*pTY/ZATV (an asterisk stands for a nick) strongly depends on the type of nearest neighbor bases X and Y flanking the nicked dinucleotide step. The maximum efficiency of the coaxial stacking was observed for the PuA*pTPy/PuATPy interface, whereas the minimum efficiency was obtained for the PyA*pTPu/PyATPu interface. A 5′‐phosphate residue in the nick enhances the coaxial stacking. In dependence on duplex structure the observed efficiency of A*T/AT coaxial stacking varied from (? 0.97 kcal/mol) for unphosphorylated TA*TA/TATA interface to three‐fold higher value (? 2.78 kcal/mol) for GA*pTT/AATC interface. 相似文献
3.
Thermodynamic parameters of coaxial stacking at complementary helix-helix interfaces GX*pYG/CZVC (X,Y=A,C,T,G;*-nick) created by contiguous oligonucleotide hybridization were determined. The data obtained were compared to the thermodynamic parameters of coaxial stacking at the interfaces CX*pYC/GZVG. Multiple linear regression analysis has revealed that the free-energy increments of interaction for the contacts GX*pYG/CZVC and CX*pYC/GZVG can be described by a set of uniform Delta G degrees(X*pY/ZV) values. The difference in the observed free-energy of the coaxial stacking between the two sets is defined by the contribution from the factors reflecting structural differences between compared DNA duplexes. 相似文献
4.
Murphy D Doyle H Eritja R Redmond G 《Journal of biomolecular structure & dynamics》2004,22(2):195-204
Three-strand oligonucleotide complexes are employed to assess the effect of base stacking and base pair mismatch on the relative thermodynamic stabilities of oligonucleotide duplexes. The melting behavior of three-strand oligonucleotide complexes incorporating nicks and gaps as well as internal single base mismatches is monitored using temperature-dependent optical absorption spectroscopy. A sequential three-state equilibrium model is used to analyze the measured melting profiles and evaluate thermodynamic parameters associated with dissociation of the complexes. The free-energy of stabilization of a nick complex compared to a gap complex due to base stacking is determined to be -1.9 kcal/mol. The influence of a mispaired base in these systems is shown to destabilize a nick complex by 3.1 kcal/mol and a gap complex by 2.8 kcal/mol, respectively. 相似文献
5.
Free energies for stacking of unpaired nucleotides (dangling ends) at the termini of oligoribonucleotide Watson-Crick helixes (DeltaG(0)37,stack) depend on sequence for 3' ends but are always small for 5' ends. Here, these free energies are correlated with stacking at helix termini in a database of 34 RNA structures determined by X-ray crystallography and NMR spectroscopy. Stacking involving GA pairs is considered separately. A base is categorized as stacked by its distance from (=4.0 A), angle with (=30 degrees), and overlap with the terminal helix base-pair. A base is unstacked if it does not satisfy one or more of these criteria.Of the 36 unpaired bases in sequences with DeltaG(0)37,stackmore favorable than -0.7 kcal/mol, 30 (83 %) are stacked on the adjacent base-pair, indicating a propensity for such sequences to stack in the 3D structure. Structures containing the strongly stacked sequence [sequence: see text] show that the amino group of C closely overlaps the carbonyl-4 of U. Thermodynamic measurement of U stacking on a 2-pyrimidinone-guanine base-pair, where the amino group of C is replaced by hydrogen, suggests that interactions with the cytosine amino group contribute approximately 0.5 kcal/mol to DeltaG(0)37,stack.For GA mismatches at helix termini, the nucleotide at the 3' helix end is always stacked, and the nucleotide at the 5' end is stacked in almost 90 % of occurrences. In available structures, non-Watson-Crick paired bases 3' to an imino-hydrogen bonded GA are also always stacked; the GA provides a large platform for favorable stacking.For the 56 sequences associated with DeltaG(0)37,stackless favorable than -0.4 kcal/mol, 19 (34 %) are stacked; these sequences have a propensity for not stacking on adjacent base-pairs. Phylogenetic conservation of weakly stacking sequences at 3' ends may be a predictor of a backbone turn. 相似文献
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8.
Kinetics of internal rotation about the C(6)-N(6) bond of N-6,N-6-dimethyladenine (M2-6A) was investigated by -1H nuclear magnetic resonance line-shape analysis of the methyl resonances (220 MHz). Rates of rotation were determined for M2-6A deuterated at N(1) and for neutral M2-6A. Activation parameters for monodeuterated M2-6A at 22 degrees are Ea = 13.8kcal/mol, log A = 12.6, incrementG++=14.9 kcal/mol, incrementH++ = 13.1 kcal/mol, incrementS++ = minus 5.8 eu; for neutral M2-6A: Ea = 15.5 kcal/mol, log A = 14.9, incrementG++ = 12.6 kcal/mol, incrementH++ = 14.9 kcal/mol, incrementS++ =7.8 eu. Vertical stacking of bases interferes with internal rotation of the dimethylamino group. 相似文献
9.
mRNA secondary structure in an open reading frame reduces translation efficiency in Bacillus subtilis. 总被引:1,自引:2,他引:1
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The structural gene for thermostable neutral protease, nprM, has only one stacking region, whose energy is -16.3 kcal/mol (-68.2 kJ/mol). Mutations for increasing (-30.8 kcal/mol [128.9 kJ/mol] and decreasing (-5.0 kcal/mol [-20.9 kJ/mol]) the energy of the stacking region were introduced in nprM on the recombinant plasmid pMK1 by using site-directed mutagenesis without any amino acid substitutions. The resultant plasmids were designated pMK2 and pMK3, respectively. The enzyme productivity of the pMK2 carrier was about 40% lower than that of pMK1, whereas the productivity of the pMK3 carrier was about 5% higher. The higher the stability of the stacking regions, the lower the enzyme productivity that was observed. mRNA concentrations were almost the same in the cells harboring these three plasmids. These results indicate that the secondary structure of mRNA reduces the translation efficiency. 相似文献
10.
The role of quaternary interactions on the stability and activity of ascorbate peroxidase. 总被引:1,自引:0,他引:1
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D. Mandelman F. P. Schwarz H. Li T. L. Poulos 《Protein science : a publication of the Protein Society》1998,7(10):2089-2098
Point mutations at the dimer interface of the homodimeric enzyme ascorbate peroxidase (APx) were constructed to assess the role of quaternary interactions in the stability and activity of APx. Analysis of the APx crystal structure shows that Glu112 forms a salt bridge with Lys20 and Arg24 of the opposing subunit near the axis of dyad symmetry between the subunits. Two point mutants, E112A and E112K, were made to determine the effects of a neutral (alanine) and repulsive (lysine) mutation on dimerization, stability, and activity. Gel filtration analysis indicated that the ratio of the monomer to dimer increased as the dimer interface interactions went from attractive to repulsive. Differential scanning calorimetry (DSC) data exhibited a decrease in both the transition temperature (Tm) and enthalpy of unfolding (deltaHc) with Tm = 58.3 +/- 0.5 degrees C, 56.0 +/- 0.8 degrees C, and 53.0 +/- 0.9 degrees C and deltaHc = 245 +/- 29 kcal/mol, 199 +/- 38 kcal/mol, and 170 +/- 25 kcal/mol for wild-type APx, E112A, and E112K, respectively. Similar changes were observed based on thermal melting curves obtained by absorption spectroscopy. No change in enzyme activity was found for the E112A mutant, and only a 25% drop in activity was observed for the E112K mutant which demonstrates that the non-Michaelis Menten kinetics of APx is not due to the APx oligomeric structure. The cryogenic crystal structures of the wild-type and mutant proteins show that mutation induced changes are limited to the dimer interface including an alteration in solvent structure. 相似文献
11.
Deirdre Murphy Hugh Doyle Ramon Eritja Gareth Redmond 《Journal of biomolecular structure & dynamics》2013,31(2):195-203
Abstract Three-strand oligonucleotide complexes are employed to assess the effect of base stacking and base pair mismatch on the relative thermodynamic stabilities of oligonucleotide duplexes. The melting behavior of three-strand oligonucleotide complexes incorporating nicks and gaps as well as internal single base mismatches is monitored using temperature-dependent optical absorption spectroscopy. A sequential three-state equilibrium model is used to analyze the measured melting profiles and evaluate thermodynamic parameters associated with dissociation of the complexes. The free-energy of stabilization of a nick complex compared to a gap complex due to base stacking is determined to be ?1.9 kcal/mol. The influence of a mispaired base in these systems is shown to destabilize a nick complex by 3.1 kcal/mol and a gap complex by 2.8 kcal/mol, respectively. 相似文献
12.
Thermodynamics of RNA internal loops with a guanosine-guanosine pair adjacent to another noncanonical pair 总被引:1,自引:0,他引:1
Thermodynamic parameters measured by optical melting are reported for formation of RNA duplexes containing tandem noncanonical pairs with at least one guanosine-guanosine (GG) pair. For selected sequences, imino proton NMR provides evidence that the desired duplex forms and that the structure of a GG pair adjacent to a noncanonical pair depends on context. A GG pair next to a different noncanonical pair is more stable than expected from measurements of adjacent GG pairs. This is likely due to an unfavorable stacking interaction between adjacent GG pairs, where areas of high negative charge probably overlap. The results suggest a model where tandem noncanonical pairs closed by two GC pairs are assigned the following free energy increments at 37 degrees C: 0.8 kcal/mol for adjacent GG pairs, 1.0 kcal/mol for GG next to UU, and -0.3 kcal/mol for all others. These values are adjusted by 0.65 kcal/mol for each closing AU pair. 相似文献
13.
E Mansini E G Oestreicher L P Ribeiro 《Archives internationales de physiologie et de biochimie》1989,97(6):447-453
Purified mitochondrial malate dehydrogenase isoenzyme (m-MDH) of Toxocara canis muscle presented maximum activity at 48 degrees C. A clear change in slope of the Arrhenius plot was observed. The energy of activation calculated for the catalytic process showed values of 3.2 kcal/mol and 10.5 kcal/mol. Thermal inactivation of m-MDH showed that it is more thermolabile than the s-isoenzyme. The inactivation of the enzyme by heat could be reduced at least in part by the addition of 0.1 mM NADH. The heat denaturation showed to be a first-order process. The rate constant (k) was calculated as being of the order of 5.28 X 10(-4) s-1 at 40 degrees C. The activation energy for the heat inactivation process was 16.45 kcal/mol between 30 degrees C and 40 degrees C and 13.79 kcal/mol between 40 degrees C and 48 degrees C. 相似文献
14.
The B-Z conformational transition in folded oligodeoxynucleotides: loop size and stability of Z-hairpins 总被引:5,自引:0,他引:5
L E Xodo G Manzini F Quadrifoglio G A van der Marel J H van Boom 《Biochemistry》1988,27(17):6327-6331
The capacity to assume a left-handed conformation and the thermodynamics of loop formation in concentrated aqueous NaClO4 have been investigated for the following palindromic sequences: d-(CGCGCGAAAAACGCGCG) (A5), d(CGCGCGTTTTTCGCGCG) (T5), d(CGCGCGTACGCGCG) (TA), and d(CGCGCGATCGCGCG) (AT). The results show that (a) each oligomer assumes a Z conformation upon exposure to increasing NaClO4 concentrations; the salt concentration at the transition midpoint is 1.8 M for both A5 and T5 and 3 and 3.5 M for TA and AT, respectively; (b) in high salt the four oligomers exist, over a wide range of nucleotide concentrations (up to 10(-3) M) and of temperature (greater than 0 degrees C), as unimolecular hairpin structures; (c) hairpins TA and AT exhibit, in buffer A, a lower thermal stability with respect to A5 and T5 (delta T about 16 degrees C), contrary to what is observed at low ionic strength; (d) on hairpin formation, the enthalpic term is about -52 kcal/mol for the two 17-mers and -38 kcal/mol for the two 14-mers, while the change in entropy is found to be around -150 eu for A5 and T5 and -115 eu for TA and AT. This thermodynamic picture suggests that a two-residue loop for TA and AT, found at low ionic strength [see preceding paper (Xodo, L.E., Manzini, G., Quadrifoglio, F., van der Marel, G.A., & van Boom, J.H. (1988) Biochemistry (preceding paper in this issue)], is substituted by a longer one including two additional residues from a missing dC.dG base pairing at the top of the stem. 相似文献
15.
Quantitative analysis of DNA secondary structure from solvent-accessible surfaces: the B- to Z-DNA transition as a model 总被引:5,自引:0,他引:5
Solvent structure and its interactions have been suggested to play a critical role in defining the conformation of polynucleotides and other macromolecules. In this work, we attempt to quantitate solvent effects on the well-studied conformational transition between right-handed B- and left-handed Z-DNA. The solvent-accessible surfaces of the hexamer sequences d(m5CG)3, d(CG)3, d(CA)3, and d(TA)3 were calculated in their B- and Z-DNA conformations. The difference in hydration free energies between the Z and the B conformations (delta delta GH(Z-B] was determined from these surfaces to be -0.494 kcal/mol for C-5 methylated d(CG), 0.228 kcal/mol for unmethylated d(CG), 0.756 kcal/mol for d(CA)-d(TG), and 0.896 kcal/mol for d(TA) dinucleotides. These delta delta GH(Z-B) values were compared to the experimental B- to Z-DNA transition energies of -0.56 kcal/mol that we measured for C-5 methylated d(CG), 0.69-1.30 kcal/mol reported for unmethylated d(CG), 1.32-1.48 kcal/mol reported for d(CA)-d(TG), and 2.3-2.4 kcal/mol for d(TA) dinucleotides. From this comparison, we found that the calculated delta delta GH(Z-B) of these dinucleotides could account for the previous observation that the dinucleotides were ordered as d(m5CG) greater than d(CG) greater than d(CA)-d(TG) greater than d(TA) in stability as Z-DNA. Furthermore, we predicted that one of the primary reasons for the inability of d(TA) sequences to form Z-DNA results from a decrease in exposed hydrophilic surfaces of adjacent base pairs due to the C-5 methyl group of thymine; thus, d(UA) dinucleotides should be more stable as Z-DNA than the analogous d(TA) dinucleotides.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
16.
Vander Meulen KA Davis JH Foster TR Record MT Butcher SE 《Journal of molecular biology》2008,384(3):702-717
Little is known about the thermodynamic forces that drive the folding pathways of higher-order RNA structure. In this study, we employ calorimetric [isothermal titration calorimetry (ITC) and differential scanning calorimetry (DSC)] and spectroscopic (NMR and UV) methods to characterize the thermodynamics of the GAAA tetraloop-receptor interaction, utilizing a previously described bivalent construct. ITC studies indicate that the bivalent interaction is enthalpy driven and highly stable, with a binding constant (K(obs)) of 5.5x10(6) M(-1) and enthalpy (DeltaH(obs)(o)) of -33.8 kcal/mol at 45 degrees C in 20 mM KCl and 2 mM MgCl(2). Thus, we derive the DeltaH(obs)(o) for a single tetraloop-receptor interaction to be -16.9 kcal/mol at these conditions. UV absorbance data indicate that an increase in base stacking quality contributes to the enthalpy of complex formation. These highly favorable thermodynamics are consistent with the known critical role for the tetraloop-receptor motif in the folding of large RNAs. Additionally, a significant heat capacity change (DeltaC(p,obs)(o)) of -0.24 kcal mol(-1) K(-1) was determined by ITC. DSC and UV-monitored thermal denaturation experiments indicate that the bivalent tetraloop-receptor construct follows a minimally five-state unfolding pathway and suggest the observed DeltaC(p,obs)(o) for the interaction results from a temperature-dependent unbound receptor RNA structure. 相似文献
17.
The reaction of horseradish peroxidase (HRP) with H2O2 has been studied in 50% v/v methanol/water over the 25.0 to -36.0 degrees C temperature range by using the low-temperature stopped-flow technique. All reactions were carried out under pseudo-first-order conditions with [H2O2] much greater than [HRP]. Arrhenius plots for the pseudo-first-order rate constant kobs were linear over the 17.6 to -36.0 degrees C temperature range studied with an activation energy of 4.8 +/- 0.5 kcal/mol. Above 0 degrees C, kobs varies linearly with peroxide concentration. However, saturation kinetics are observed below -16.0 degrees C, indicating that there is at least one reversible elementary step in this reaction. Double-reciprocal plots at -26.0 degrees C at pH* 7.3 for the reaction give kappa max(obs) = 163 s-1 and KM = 0.190 mM. Rapid-scan optical studies carried out at -35.0 degrees C with [H2O2] much greater than KM reveal the presence of a transient intermediate referred to as compound 0 whose conversion to compound I is rate limiting. The Soret region of the optical spectrum of compound 0 resembles that of a "hyperporphyrin" with prominent bands near 330 and 410 nm. The temperature dependencies of kappa max(obs) and KM have been measured over the -16.0 to -26.0 degrees C range and give an activation energy for kappa max(obs) of 1.6 +/- 0.7 kcal/mol and an enthalpy of formation for compound 0 of 4.0 +/- 0.7 kcal/mol. 相似文献
18.
Ruan Q Liu T Kolbanovskiy A Liu Y Ren J Skorvaga M Zou Y Lader J Malkani B Amin S Van Houten B Geacintov NE 《Biochemistry》2007,46(23):7006-7015
The influence of DNA base sequence context on the removal of a bulky benzo[a]pyrene diol epoxide-guanine adduct, (+)-trans-B[a]P-N2-dG (G*), by UvrABC nuclease from the thermophilic organism Bacillus caldotenax was investigated. The lesion was flanked by either T or C in otherwise identical complementary 43-mer duplexes (TG*T or CG*C, respectively). It was reported earlier that in the CG*C context, a dominant minor groove adduct structure was observed by NMR methods with all Watson-Crick base pairs intact, and the duplex exhibited a rigid bend. In contrast, in the TG*T context, a highly flexible bend was observed, base pairing at G*, and two 5'-base pairs flanking the adduct were impaired, and multiple solvent-accessible adduct conformations were observed. The TG*T-43-mer duplexes are incised with consistently greater efficiency by UvrABC proteins from B. caldotenax by a factor of 2.3 +/- 0.3. The rates of incisions increase with increasing temperature and are characterized by linear Arrhenius plots with activation energies of 27.0 +/- 1.5 and 23.4 +/- 1.0 kcal/mol for CG*C and TG*T duplexes, respectively. These values reflect the thermophilic characteristics of the UVrABC nuclease complex and the contributions of the different DNA substrates to the overall activation energies. These effects are consistent with base sequence context-dependent differences in structural disorder engendered by a loss of local base stacking interactions and Watson-Crick base pairing in the immediate vicinity of the lesions in the TG*T duplexes. The local weakening of base pairing interactions constitutes a recognition element of the UvrABC nucleotide excision repair apparatus. 相似文献
19.
Several models have been proposed to explain the high temperatures required to denature enzymes from thermophilic organisms; some involve greater maximum thermodynamic stability for the thermophile, and others do not. To test these models, we reversibly melted two analogous protein domains in a two-state manner. E2cd is the isolated catalytic domain of cellulase E2 from the thermophile Thermomonospora fusca. CenAP30 is the analogous domain of the cellulase CenA from the mesophile Cellulomonas fimi. When reversibly denatured in a common buffer, the thermophilic enzyme E2cd had a temperature of melting (Tm) of 72.2 degrees C, a van't Hoff enthalpy of unfolding (DeltaHVH) of 190 kcal/mol, and an entropy of unfolding (DeltaSu) of 0.55 kcal/(mol*K); the mesophilic enzyme CenAP30 had a Tm of 56.4 degrees C, a DeltaHVH of 107 kcal/mol, and a DeltaSu of 0. 32 kcal/(mol*K). The higher DeltaHVH and DeltaSu values for E2cd suggest that its free energy of unfolding (DeltaGu) has a steeper dependence on temperature at the Tm than CenAP30. This result supports models that predict a greater maximum thermodynamic stability for thermophilic enzymes than for their mesophilic counterparts. This was further explored by urea denaturation. Under reducing conditions at 30 degrees C, E2cd had a concentration of melting (Cm) of 5.2 M and a DeltaGu of 11.2 kcal/mol; CenAP30 had a Cm of 2.6 M and a DeltaGu of 4.3 kcal/mol. Under nonreducing conditions, the Cm and DeltaGu of CenAP30 were increased to 4.5 M and 10.8 kcal/mol at 30 degrees C; the Cm for E2cd was increased to at least 7.4 M at 32 degrees C. We were unable to determine a DeltaGu value for E2cd under nonreducing conditions due to problems with reversibility. These data suggest that E2cd attains its greater thermal stability (DeltaTm = 15.8 degrees C) through a greater thermodynamic stability (DeltaDeltaGu = 6.9 kcal/mol) compared to its mesophilic analogue CenAP30. 相似文献
20.
Meneni SR Shell SM Gao L Jurecka P Lee W Sponer J Zou Y Chiarelli MP Cho BP 《Biochemistry》2007,46(40):11263-11278
A systematic spectroscopic and computational study was conducted in order to probe the influence of base sequences on stacked (S) versus B-type (B) conformational heterogeneity induced by the major dG adduct derived from the model carcinogen 7-fluoro-2-aminofluorene (FAF). We prepared and characterized eight 12-mer DNA duplexes (-AG*N- series, d[CTTCTAG*NCCTC]; -CG*N- series, d[CTTCTCG*NCCTC]), in which the central guanines (G*) were site-specifically modified with FAF with varying flanking bases (N = G, A, C, T). S/B heterogeneity was examined by CD, UV, and dynamic 19F NMR spectroscopy. All the modified duplexes studied followed a typical dynamic exchange between the S and B conformers in a sequence dependent manner. Specifically, purine bases at the 3'-flanking site promoted the S conformation (G > A > C > T). Simulation analysis showed that the S/B energy barriers were in the 14-16 kcal/mol range. The correlation times (tau = 1/kappa) were found to be in the millisecond range at 20 degrees C. The van der Waals energy force field calculations indicated the importance of the stacking interaction between the carcinogen and neighboring base pairs. Quantum mechanics calculations showed the existence of correlations between the total interaction energies (including electrostatic and solvation effects) and the S/B population ratios. The S/B equilibrium seems to modulate the efficiency of Escherichia coli UvrABC-based nucleotide excision repair in a conformation-specific manner: i.e., greater repair susceptibility for the S over B conformation and for the -AG*N- over the -CG*N- series. The results indicate a novel structure-function relationship, which provides insights into how bulky DNA adducts are accommodated by UvrABC proteins. 相似文献