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1.
Structures of mismatched base pairs in DNA and their recognition by the Escherichia coli mismatch repair system. 总被引:8,自引:1,他引:7
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G V Fazakerley E Quignard A Woisard W Guschlbauer G A van der Marel J H van Boom M Jones M Radman 《The EMBO journal》1986,5(13):3697-3703
The Escherichia coli mismatch repair system does not recognize and/or repair all mismatched base pairs with equal efficiency: whereas transition mismatches (G X T and A X C) are well repaired, the repair of some transversion mismatches (e.g. A X G or C X T) appears to depend on their position in heteroduplex DNA of phage lambda. Undecamers were synthesized and annealed to form heteroduplexes with a single base-pair mismatch in the centre and with the five base pairs flanking each side corresponding to either repaired or unrepaired heteroduplexes of lambda DNA. Nuclear magnetic resonance (n.m.r.) studies show that a G X A mismatch gives rise to an equilibrium between fully helical and a looped-out structure. In the unrepaired G X A mismatch duplex the latter predominates, while the helical structure is predominant in the case of repaired G X A and G X T mismatches. It appears that the E. coli mismatch repair enzymes recognize and repair intrahelical mismatched bases, but not the extrahelical bases in the looped-out structures. 相似文献
2.
Heteroduplex DNA correction in Saccharomyces cerevisiae is mismatch specific and requires functional PMS genes. 总被引:35,自引:13,他引:22
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In vitro-constructed heteroduplex DNAs with defined mismatches were corrected in Saccharomyces cerevisiae cells with efficiencies that were dependent on the mismatch. Single-nucleotide loops were repaired very efficiently; the base/base mismatches G/T, A/C, G/G, A/G, G/A, A/A, T/T, T/C, and C/T were repaired with a high to intermediate efficiency. The mismatch C/C and a 38-nucleotide loop were corrected with low efficiency. This substrate specificity pattern resembles that found in Escherichia coli and Streptococcus pneumoniae, suggesting an evolutionary relationship of DNA mismatch repair in pro- and eucaryotes. Repair of the listed mismatches was severely impaired in the putative S. cerevisiae DNA mismatch repair mutants pms1 and pms2. Low-efficiency repair also characterized pms3 strains, except that correction of single-nucleotide loops occurred with an efficiency close to that of PMS wild-type strains. A close correlation was found between the repair efficiencies determined in this study and the observed postmeiotic segregation frequencies of alleles with known DNA sequence. This suggests an involvement of DNA mismatch repair in recombination and gene conversion in S. cerevisiae. 相似文献
3.
W N Hunter T Brown G Kneale N N Anand D Rabinovich O Kennard 《The Journal of biological chemistry》1987,262(21):9962-9970
The structure of the deoxyoligomer d(C-G-C-G-A-A-T-T-T-G-C-G) was determined at 2.5-A resolution by single crystal x-ray diffraction techniques. The final R factor is 18% with the location of 71 water molecules. The oligomer crystallizes in a B-DNA-type conformation, with two strands interacting to form a dodecamer duplex. The double helix consists of four A X T and six G X C Watson-Crick base pairs and two G X T mismatches. The G X T pairs adopt a "wobble" structure with the thymine projecting into the major groove and the guanine into the minor groove. The mispairs are accommodated in the normal double helix by small adjustments in the conformation of the sugar phosphate backbone. A comparison with the isomorphous parent compound containing only Watson-Crick base pairs shows that any changes in the structure induced by the presence of G X T mispairs are highly localized. The global conformation of the duplex is conserved. The G X T mismatch has already been studied by x-ray techniques in A and Z helices where similar results were found. The geometry of the mispair is essentially identical in all structures so far examined, irrespective of the DNA conformation. The hydration is also similar with solvent molecules bridging the functional groups of the bases via hydrogen bonds. Hydration may be an important factor in stabilizing G X T mismatches. A characteristic of Watson-Crick paired A X T and G X C bases is the pseudo 2-fold symmetry axis in the plane of the base pairs. The G X T wobble base pair is pronouncedly asymmetric. This asymmetry, coupled with the disposition of functional groups in the major and minor grooves, provides a number of features which may contribute to the recognition of the mismatch by repair enzymes. 相似文献
4.
Base mismatch-specific endonuclease activity in extracts from Saccharomyces cerevisiae. 总被引:1,自引:0,他引:1
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An endonuclease activity (called MS-nicking) for all possible base mismatches has been detected in the extracts of yeast, Saccharomyces cerevisiae. DNAs with twelve possible base mismatches at one defined position are cleaved at different efficiencies. DNA fragments with A/G, G/A, T/G, G/T, G/G, or A/A mismatches are nicked with greater efficiencies than C/T, T/C, C/A, and C/C. DNA with an A/C or T/T mismatch is nicked with an intermediate efficiency. The MS-nicking is only on one particular DNA strand, and this strand disparity is not controlled by methylation, strand break, or nature of the mismatch. The nicks have been mapped at 2-3 places at second, third, and fourth phosphodiester bonds 5' to the mispaired base; from the time course study, the fourth phosphodiester bond probably is the primary incision site. This activity may be involved in mismatch repair during genetic recombination. 相似文献
5.
Repair of Single Base-Pair Transversion Mismatches of Escherichia Coli in Vitro: Correction of Certain a/G Mismatches Is Independent of Dam Methylation and Host Muthls Gene Functions 总被引:17,自引:0,他引:17
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Six different base-pair transversion mismatches are repaired with different efficiencies in an in vitro mismatch repair system. In particular, the T/T and C/C mismatches appear to be less efficiently repaired than the A/A and G/G mismatches. Four A/G and four C/T mismatches at different positions are repaired to different extents. One of the A/G mismatches is repaired equally efficiently when DNA heteroduplexes are fully methylated or hemi-methylated at the d(GATC) sequences. This type of mismatch repair appears to be unidirectional with A to C conversion by acting at A/G mispairs to restore the C/G pairs. This methylation-independent correction is not controlled by the mutH, mutL, mutS, uvrE, uvrB, phr, recA, recF, and recJ gene products. The independence of the transversion mismatch repair of these genes and methylation distinguishes this from the known mismatch repair pathways. 相似文献
6.
M Radman C Dohet M Jones M P Doutriaux F Laengle-Rouault G Maenhaut-Michel R Wagner 《Biochimie》1985,67(7-8):745-752
Bacteriophage lambda and phi X 174 DNAs, carrying sequenced mutations, have been used to construct in vitro defined species of heteroduplex DNA. Such heteroduplex DNAs were introduced by transfection, as single copies, into E. coli host cells. The progeny of individual heteroduplex molecules from each infective center was analyzed. The effect of the presence of GATC sequences (phi X 174 system) and of their methylation (lambda system) was tested. The following conclusions can be drawn: some mismatched base pairs trigger the process of mismatch repair, causing a localized strand-to-strand information transfer in heteroduplex DNA: transition mismatches G:T and A:C are efficiently repaired, whereas the six transversion mismatches are not always readily recognized and/or repaired. The recognition of transversion mismatches appears to depend on the neighbouring nucleotide sequence; single unpaired bases (frameshift mutation "mismatches") are recognized and repaired, some equally efficiently on both strands (longer and shorter), some more efficiently on the shorter (-1) strand; large non-homologies (about 800 bases) are not repaired by the Mut H, L, S, U system, but some other process repairs the non-homology with a relatively low efficiency; full methylation of GATC sequences inhibits mismatch repair on the methylated strand: this is the chemical basis of strand discrimination (old/new) in mismatch correction; unmethylated GATC sequences appear to improve mismatch repair of a G:T mismatch in phi X 174 DNA, but there may be some residual mismatch repair in GATC-free phi X 174, at least for some mismatches.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
7.
Repair of Single- and Multiple-Substitution Mismatches during Recombination in Streptococcus Pneumoniae 总被引:10,自引:0,他引:10
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The use as genetic markers, during transformation of Streptococcus pneumoniae, of 19 sequences differing from wild type, located throughout the amiA locus, enabled us to examine the fate of 24 single- and 11 multiple-mismatches during recombination. Tentative mismatch ranking as a function of decreasing repair efficiency by the Hex mismatch repair system is G/T = A/C = G/G (maximum repair: 90-95%) greater than C/T (mostly 75 to 90% repair) greater than A/A (from 50 to 90% repair) greater than T/T (50-65% repair) greater than A/G (from 0 to 20% repair) greater than C/C. No indication of correction of the latter has been obtained. Over the limited number of samples examined, we observed no influence of the base composition of the surrounding sequence on correction efficiency for both transition mismatches and for G/G and C/C. Variations in the surrounding sequence affect repair of A/G and C/T, and, even more strongly, of A/A and T/T. No simple correlation to the G:C content of the surrounding sequence is apparent from our results, in contrast to the conclusion drawn for the Mut mismatch repair system of Escherichia coli. Examination of the fate of multiple mismatches suggests that C/C may sometimes impede recognition of otherwise corrected mismatches. 相似文献
8.
Mismatch repair proteins collaborate with methyltransferases in the repair of O(6)-methylguanine 总被引:1,自引:0,他引:1
DNA repair is essential for combatting the adverse effects of damage to the genome. One example of base damage is O(6)-methylguanine (O(6)mG), which stably pairs with thymine during replication and thereby creates a promutagenic O(6)mG:T mismatch. This mismatch has also been linked with cellular toxicity. Therefore, in the absence of repair, O(6)mG:T mismatches can lead to cell death or result in G:C-->A:T transition mutations upon the next round of replication. Cysteine thiolate residues on the Ada and Ogt methyltransferase (MTase) proteins directly reverse the O(6)mG base damage to yield guanine. When a cytosine is opposite the lesion, MTase repair restores a normal G:C pairing. However, if replication past the lesion has produced an O(6)mG:T mismatch, MTase conversion to a G:T mispair must still undergo correction to avoid mutation. Two mismatch repair pathways in E. coli that convert G:T mispairs to native G:C pairings are methyl-directed mismatch repair (MMR) and very short patch repair (VSPR). This work examined the possible roles that proteins in these pathways play in coordination with the canonical MTase repair of O(6)mG:T mismatches. The possibility of this repair network was analyzed by probing the efficiency of MTase repair of a single O(6)mG residue in cells deficient in individual mismatch repair proteins (Dam, MutH, MutS, MutL, or Vsr). We found that MTase repair in cells deficient in Dam or MutH showed wild-type levels of MTase repair. In contrast, cells lacking any of the VSPR proteins MutS, MutL, or Vsr showed a decrease in repair of O(6)mG by the Ada and Ogt MTases. Evidence is presented that the VSPR pathway positively influences MTase repair of O(6)mG:T mismatches, and assists the efficiency of restoring these mismatches to native G:C base pairs. 相似文献
9.
Repair of specific base pair mismatches formed during meiotic recombination in the yeast Saccharomyces cerevisiae. 总被引:13,自引:6,他引:7
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Heteroduplexes formed between DNA strands derived from different homologous chromosomes are an intermediate in meiotic crossing over in the yeast Saccharomyces cerevisiae and other eucaryotes. A heteroduplex formed between wild-type and mutant genes will contain a base pair mismatch; failure to repair this mismatch will lead to postmeiotic segregation (PMS). By analyzing the frequency of PMS for various mutant alleles in the yeast HIS4 gene, we showed that C/C mismatches were inefficiently repaired relative to all other point mismatches. These other mismatches (G/G, G/A, T/T, A/A, T/C, C/A, A/A, and T/G) were repaired with approximately the same efficiency. We found that in spores with unrepaired mismatches in heteroduplexes, the nontranscribed strand of the HIS4 gene was more frequently donated than the transcribed strand. In addition, the direction of repair for certain mismatches was nonrandom. 相似文献
10.
Structural studies of DNA fragments: the G.T wobble base pair in A, B and Z DNA; the G.A base pair in B-DNA 总被引:10,自引:0,他引:10
O Kennard 《Journal of biomolecular structure & dynamics》1985,3(2):205-226
The crystal structures of five double helical DNA fragments containing non-Watson-Crick complementary base pairs are reviewed. They comprise four fragments containing G.T base pairs: two deoxyoctamers d(GGGGCTCC) and d(GGGGTCCC) which crystallise as A type helices; a deoxydodecamer d(CGCGAATTTGCG) which crystallises in the B-DNA conformation; and the deoxyhexamer d(TGCGCG), which crystallises as a Z-DNA helix. In all four duplexes the G and T bases form wobble base pairs, with bases in the major tautomer forms and hydrogen bonds linking N1 of G with O2 of T and O6 of G with N3 of T. The X-ray analyses establish that the G.T wobble base pair can be accommodated in the A, B or Z double helix with minimal distortion of the global conformation. There are, however, changes in base stacking in the neighbourhood of the mismatched bases. The fifth structure, d(CGCGAATTAGCG), contains the purine purine mismatch G.A where G is in the anti and A in the syn conformation. The results represent the first direct structure determinations of base pair mismatches in DNA fragments and are discussed in relation to the fidelity of replication and mismatch recognition. 相似文献
11.
Different base/base mispairs are corrected with different efficiencies and specificities in monkey kidney cells 总被引:47,自引:0,他引:47
Mismatches arise during recombination, as errors of DNA replication, and from deamination of 5-methylcytosine to thymine. We determined the efficiency and specificity of mismatch correction in simian cells. Analysis of plaques, obtained after transfection with SV40 DNA molecules harboring a single mispair in a defined orientation within the intron of the large T antigen gene, revealed that all types of base/base mispairs were corrected, albeit with different efficiencies and specificities. Heterogeneous mispairs G/T, A/C, C/T, and A/G, corrected with 96%, 78%, 72%, and 39% efficiencies, respectively, tended to be corrected to G/C. Homogeneous mispairs G/C, C/C, A/A, and T/T were corrected with 92%, 66%, 58%, and 39% efficiencies, respectively, and repair bias was influenced by mismatch flanking sequences. 相似文献
12.
Meiotic Mismatch Repair Quantified on the Basis of Segregation Patterns in Schizosaccharomyces Pombe 总被引:5,自引:2,他引:3
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Hybrid DNA with mismatched base pairs is a central intermediate of meiotic recombination. Mismatch repair leads either to restoration or conversion, while failure of repair results in post-meiotic segregation (PMS). The behavior of three G to C transversions in one-factor crosses with the wild-type alleles is studied in Schizosaccharomyces pombe. They lead to C/C and G/G mismatches and are compared with closely linked mutations yielding other mismatches. A method is presented for the detection of PMS in random spores. The procedure yields accurate PMS frequencies as shown by comparison with tetrad data. A scheme is presented for the calculation of the frequency of hybrid DNA formation and the efficiency of mismatch repair. The efficiency of C/C repair in S. pombe is calculated to be about 70%. Other mismatches are repaired with close to 100% efficiency. These results are compared with data published on mutations in Saccharomyces cerevisiae and Ascobolus immersus. This study forms the basis for the detailed analysis of the marker effects caused by G to C transversions in two-factor crosses. 相似文献
13.
Marker Effects of G to C Transversions on Intragenic Recombination and Mismatch Repair in Schizosaccharomyces Pombe 总被引:5,自引:2,他引:3
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G to C transversion mutations show very strong allele-specific marker effects on the frequency of wild-type recombinants in intragenic two-factor crosses. Here we present a detailed study of the marker effect of one representative, the ade6-M387 mutation of Schizosaccharomyces pombe. Crosses of M387 with other mutations at varying distance reveal highly increased prototroph frequencies in comparison with the C to T transition mutation ade6-51 (control without any known marker effect) located four nucleotides from M387. The marker effect of M387 is strongest (>40-fold) for crosses with mutations less than 15 nucleotides from M387. It decreases to an intermediate level (5-10-fold) in crosses with mutations located 25-150 base pairs from M387/51 and is very low in crosses with mutations beyond 200 base pairs. On the basis of these results and the quantitation of the low efficiency of C/C mismatch repair presented in the accompanying publication we propose the existence of at least two different types of mechanisms for base mismatch repair in fission yeast. The major system is suggested to recognize all base mismatches except C/C with high efficiency and to generate long excision tracts (approximately 100 nucleotides unidirectionally). The minor system is proposed to recognize all base mismatches including C/C with low and variable efficiency and to have short excision tracts (approximately 10 nucleotides unidirectionally). We estimate from the M387 marker effect that the minor system accounts for approximately 1-8% repair of non-C/C mismatches (depending on the nature of the mutation) in fission yeast meiosis. 相似文献
14.
Structural features of pyrimidine.pyrimidine mismatches in the interior of oligonucleotide duplexes have been investigated by high resolution two-dimensional proton nuclear magnetic resonance (n.m.r.) spectroscopy. These studies were conducted on the self-complementary d(C-G-C-T-A-G-C-T-T-G-C-G) duplex (designated T.T 12-mer) and the self-complementary d(C-G-C-C-A-G-C-T-C-G-C-G) duplex (designated C.C 12-mer) containing T.T and C.C pairs located at identical positions four base-pairs from either end of the duplex. Proton n.m.r. studies on the T.T 12-mer duplex were undertaken in the neutral pH range, while studies on the C.C 12-mer duplex were recorded at acidic pH. The proton spectra narrowed considerably on lowering the pH below neutrality for the C.C 12-mer duplex. Two-dimensional nuclear Overhauser enhancement spectroscopy (NOESY) data sets have been recorded on the T.T 12-mer and C.C 12-mer duplexes in high salt H2O and D2O solution. The magnitude of the NOE crosspeaks and the directionality of the NOE connectivities demonstrate that both duplexes are right-handed with all bases, including those at the mismatch site, adopting an anti configuration about the glycosidic bond. The observed base and sugar proton chemical shifts suggest structural similarities for the trinucleotide segments centered about the T.T and C.C mismatches. A NOE is detected between the resolved imino protons of T4 and T9 at the mismatch site, consistent with formation of a stacked "wobble" T4(anti).T9(anti) pair in the T.T 12-mer duplex. A comparison of the imino proton chemical shift and NOE data suggests that the imino-carbonyl hydrogen bonds in the wobble T.T mismatch are weaker than the corresponding imino-carbonyl hydrogen bonds in the wobble G.T mismatch. The 4-amino protons of C4 and C9 at the mismatch site in the C.C 12-mer duplex do not exhibit the pattern of hydrogen-bonded and exposed protons separated by approximately 1.5 parts per million characteristic of cytidine amino protons involved in Watson-Crick G.C pairing. The experimental data are insufficient to differentiate between wobble C(anti).C+(anti) and other pairing possibilities for the mismatch in the C.C 12-mer duplex at acidic pH. 相似文献
15.
Olga Kennard 《Journal of biomolecular structure & dynamics》2013,31(2):205-226
Abstract The crystal structures of five double helical DNA fragments containing non-Watson-Crick complementary base pairs are reviewed. They comprise four fragments containing G·T base pairs: two deoxyoctamers d(GGGGCTCC) and d(GGGGTCCC) which crystallise as A type helices; a deoxydodecamer d(CGCGAATTTGCG) which crystallises in the B-DNA conformation; and the deoxyhexamer d(TGCGCG), which crystallises as a Z-DNA helix. In all four duplexes the G and T bases form wobble base pairs, with bases in the major tautomer forms and hydrogen bonds linking N1 of G with 02 of T and 06 of G with N3 of T. The X-ray analyses establish that the G·T wobble base pair can be accommodated in the A, B or Z double helix with minimal distortion of the global conformation. There are, however, changes in base stacking in the neighbourhood of the mismatched bases. The fifth structure, d(CGCGAATTAGCG), contains the purine purine mismatch G·A where G is in the anti and A in the syn conformation. The results represent the first direct structure determinations of base pair mismatches in DNA fragments and are discussed in relation to the fidelity of replication and mismatch recognition. 相似文献
16.
DNA sequence analysis was undertaken to investigate the structural basis of mutations showing different integration efficiencies in Streptococcus pneumoniae. Wild type, mutant and revertant sequences at two sites in the amiA locus were determined. It appears that markers which transform efficiently or inefficiently can result from single base pair changes. A low efficiency (LE) marker corresponds to a C:G to T:A change and a high efficiency (HE) marker to a G:C to T:A change. In the latter case, two mismatches, G/A and T/C, can exist at the heteroduplex stage in transformation; only T/C appears to be recognized by the hex system which controls transforming efficiencies in pneumococcus. Each of the recognized mismatches, T/G and C/A, which result from transitional change, and T/C appears to involve at least one pyrimidine. It is proposed that the mismatch repair system of S. pneumoniae is directed against mismatched pyrimidines. DNA sequence analysis also reveals that short deletions (33 or 34 bases long) behave as very high efficiency markers, confirming that deletions are not recognized by the hex system. 相似文献
17.
We have studied the formation of DNA triple helices at target sites that contain mismatches in the duplex target. Fluorescence melting studies were used to examine a series of parallel triple helices that contain all 64 N.XZ triplet combinations at the centre (where N, X and Z are each of the four natural DNA bases in turn). Similar experiments were also performed with N=bis-amino-U (BAU) (for stable recognition of AT base pairs) and N=S (for recognition of TA inversions). We find that the introduction of a duplex mismatch destabilises the C+.GZ, T.AZ and G.TZ triplets. A similar effect is seen with BAU.AZ triplets. In contrast, other base combinations, based on non-standard triplets such as C.AZ, T.TZ, G.CZ and A.CZ are stabilised by the presence of a duplex mismatch. In each case S binds to sites containing duplex mismatches better than the corresponding Watson-Crick base pairs. 相似文献
18.
Influence of nearest neighbor sequence on the stability of base pair mismatches in long DNA; determination by temperature-gradient gel electrophoresis. 总被引:9,自引:3,他引:6
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Temperature-gradient gel electrophoresis (TGGE) was employed to determine the thermal stabilities of 48 DNA fragments that differ by single base pair mismatches. The approach provides a rapid way for studying how specific base mismatches effect the stability of a long DNA fragment. Homologous 373 bp DNA fragments differing by single base pair substitutions in their first melting domain were employed. Heteroduplexes were formed by melting and reannealing pairs of DNAs, one of which was 32P-labeled on its 5'-end. Product DNAs were separated based on their thermal stability by parallel and perpendicular temperature-gradient gel electrophoresis. The order of stability was determined for all common base pairs and mismatched bases in four different nearest neighbor environments; d(GXT).d(AYC), d(GXG).d(CYC), d(CXA).d(TYG), and d(TXT).d(AYA) with X,Y = A, T, C, or G. DNA fragments containing a single mismatch were destabilized by 1 to 5 degrees C with respect to homologous DNAs with complete Watson-Crick base pairing. Both the bases at the mismatch site and neighboring stacking interactions influence the destabilization caused by a mismatch. G.T, G.G and G.A mismatches were always among the most stable mismatches for all nearest neighbor environments examined. Purine.purine mismatches were generally more stable than pyrimidine.pyrimidine mispairs. Our results are in very good agreement with data where available from solution studies of short DNA oligomers. 相似文献
19.
Sung Jae Cho Joon-Hwa Lee Byong-Seok Choi 《Archives of biochemistry and biophysics》2010,501(2):201-206
In Escherichia coli, the very short patch (VSP) repair system is a major pathway for removal of T·G mismatches in Dcm target sequences. In the VSP repair pathway, the very short patch repair (Vsr) endonuclease selectively recognizes a T·G mismatch in Dcm target sequences and hydrolyzes the 5′-phosphate group of the mismatched thymine. The hydrogen exchange NMR studies here revealed that the T5·G18 mismatch in the Dcm target sequence significantly stabilizes own base pair but destabilizes the two neighboring G4·C19 and A6·T17 base pairs compare to other T·G mismatches. These unusual patterns of base pair stability in the Dcm target sequence can explain how the Vsr endonuclease specifically recognizes the mismatched Dcm target sequence and intercalates into the DNA. 相似文献
20.
Repair of a Mismatch Is Influenced by the Base Composition of the Surrounding Nucleotide Sequence 总被引:21,自引:2,他引:19
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Heteroduplexes with single base pair mismatches of known sequence were prepared by annealing separated strands of bacteriophage lambda DNA and used to transfect Escherichia coli. A series of transition (G:T and A:C) and transversion (G:A and C:T) mismatches located throughout most of the bacteriophage lambda cI gene has been examined. The results suggest that the transition mismatches are generally better repaired than the transversion mismatches and that, at least for the transversion mismatches studied, repair efficiency increases with increasing G:C content in the neighboring nucleotide sequence. This specificity of the E. coli mismatch repair system can account, in part, for the similar frequencies of base substitution mutations throughout the E. coli genome. 相似文献