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Results for “Multiprotein Complexes”

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PubMed1995

115 kDa protein from Xenopus laevis embryos recognized by antibodies directed against the Xenopus homeoprotein XIHbox 1.

Using antibodies against homeoprotein XIHbox 1 from Xenopus laevis, we have detected a new embryonic protein with a much larger molecular weight, 115 kDa. Antibodies fractionated according to their affinity for 3 different domains of the XIHbox 1 protein were used to show that this new protein is related to the C-terminal region of XIHbox 1 protein, downstream from the homeodomain. By immunohistochemistry, the protein was shown to be localized in nuclei of embryonic cells. On SDS-polyacrylamide gels, the 115 kDa protein appears as a set of closely spaced bands whose pattern varies with the stage of development and with the parental origin of the embryos. The protein could be extracted from embryos in a multiprotein complex of approximately 600 kDa. In contrast, the 18 and 27 kDa proteins predicted from the sequence of cloned cDNA to be transcribed and translated from the XIHbox 1 gene could not be detected, suggesting that they are rare or unstable in embryos. These data suggest that the new protein is involved in the development of Xenopus embryos, with a function possibly related to that of the homeoprotein XIHbox 1.

Animals↗

PubMed1994

Deoxyribonucleotides are maintained at normal levels in a yeast thioredoxin mutant defective in DNA synthesis.

Deletion of both thioredoxin genes TRX1 and TRX2 of Saccharomyces cerevisiae reduces the rate of DNA replication. This observation, originally determined by flow cytometry, was confirmed by radiochemical labeling of synchronized cultures. Since thioredoxin is a hydrogen donor to ribonucleotide reductase, a priori the inhibition of DNA synthesis was predicted to be caused by a reduction in the deoxyribonucleotide pools. However, the levels of TTP, dCTP, dATP, and dGTP were either unchanged or slightly greater in the thioredoxin mutant (3.2, 0.91, 1.4, and 1.21 pmol/10(6) cells, respectively) versus the wild-type culture (2.5, 0.91, 1.0, and 0.68 pmol/10(6) cells, respectively). An impact on ribonucleotide reduction was seen by an increased accumulation of RNR1 and RNR2 transcripts in the thioredoxin mutant (4.3- and 6.8-fold, respectively). Increased RNR expression did not reflect a general response of the DNA replication machinery. POL1 (DNA polymerase I) and CDC8 (thymidylate kinase) transcription were unaltered, while histone H2B transcripts actually decreased by half. Two alternative models incorporating these results are discussed. One suggests that thioredoxin reduces a multiprotein complex channeling nucleotides to the replication apparatus. The second proposes that thioredoxin regulates the tempo of DNA replication directly by activating a component of the replication machinery.

DNA Replication↗

PubMed1993

Immunopurification of yeast TATA-binding protein and associated factors. Presence of transcription factor IIIB transcriptional activity.

The TATA-binding proteins (TBP) from both human and Drosophila have been shown to exist in various distinct multiprotein complexes that are required, respectively, for transcription by all three RNA polymerases. In contrast, in vitro biochemical analyses have suggested that yeast TBP exists as a monomeric 27-kDa protein free in solution. We have examined the oligomerization state of yeast TBP and report here that yeast TBP, like human and Drosophila TBPs, is also stably associated with other proteins in vitro. Using anti-TBP antibodies we have immunopurified yeast TBP and associated factors (TBP-associated factors or TAFs). When this fraction was analyzed by SDS-polyacrylamide gel electrophoresis, polypeptides of approximate relative molecular size ranging from 170 to 60 kDa are prominently represented. Immunoblot analysis revealed that one of these TAFs, TAF70, corresponds to BRF1/TDS4/PCF4, a subunit of transcription factor (TF) IIIB. Furthermore, this highly purified TAF fraction can reconstitute polymerase III transcription when supplemented with purified RNA polymerase III and TFIIIC. Our data indicate that our TAF fraction contains TFIIIB transcription factor activity and that all the subunits of yeast TFIIIB are stably complexed with TBP.

Base Sequence↗

PubMedDate not supplied

DNA lesion-recognizing proteins and the p53 connection.

A great deal of the energy and time of a cell is invested in DNA repair activities. The first step in DNA repair pathways is recognition of the lesion on the DNA. The classical lesion-recognizing proteins interact with other repair proteins to form multiprotein complexes most notable of which are those that function in Nucleotide Excision Repair (NER). Proteins involved in lesion recognition include HMG1 and 2 recognizing cisplatin adducts but also maintaining active nucleosome structures and interacting with loops in cruciforms; HMG-box nuclear proteins; XPA and XPC lacking in xeroderma pigmentosum patients and involved in lesion recognition during NER; p53 recognizing strand breaks and insertion/deletion mismatches and causing arrest in the cell cycle; MSH2 mismatch repair protein identified as the human colon cancer gene product; and others including the transcription factor YB-1 that binds to depurinated DNA with a higher affinity compared with undamaged DNA. Other type of lesion-recognizing proteins are also repair enzymes like the O(6)-methylguanine-DNA methyltransferase and DNA glycosylases. Lesion recognition is an important process and might be the rate-limiting step in the overall repair process.

Animals↗

PubMed1996

A role for a p21-E2F interaction during senescence arrest of normal human fibroblasts.

The family of E2F transcription factors forms different multiprotein complexes with cell cycle regulatory proteins to control the expression of genes important in cell proliferation. In this study, we identified four distinct E2F complexes present in aged and senescent normal, human diploid fibroblasts. Two appeared to be identical to the previously described G1-specific p130 and Rb-E2F complexes present in young G0-arrested cells. The other two were novel E2F complexes that contained the cyclin-dependent kinase inhibitor p21 (cip1/WAF1/Sdi1/CAP20/PIC1) complexed with Rb/CDK2/cyclin E or with the Rb-related p107/CDK2/ cyclin D. These p21-E2F complexes, while present in young G1 cells at very low levels, were elevated in senescent cells. The p21 containing E2F complexes were not detected during the S-phase in young cells. The DNA-binding stability of the p21 complexes was approximately 10 times greater than the stability of any other E2F complex or uncomplexed E2F. Addition of purified p21 protein to the S-phase-specific cyclin A/ CDK2-p107-E2F complex from young cells dissociated cyclin A and CDK2 from p107/E2F, suggesting an additional novel function for p21. Finally, expression of p21 specifically inhibited transcription from an E2F-dependent promoter but had no effect on a mutant E2 promoter. In addition to its inhibition of CDK enzymes and proliferating cell nuclear antigen function in DNA replication, these studies reveal a novel mechanism by which p21 mediates growth arrest: direct interaction with E2F complexes and negative regulation of E2F transcription factor activity.

CDC2-CDC28 Kinases↗

PubMed1995

A human autoantibody recognizing nuclear matrix-associated nuclear protein localized in dot structures.

A human autoimmune serum is used to characterize a protein which is located within a dot-like structure of the interphase nucleus. The dots are located in the nucleoplasma outside the nucleoli. The dot-like structure could be observed on a variety of human substrates (HEp-2, HepG2, HeLa, Molt-4, WI-38 cell lines, peripheral blood lymphocytes), mouse cell lines and tissues (3T3, L929, spleen sections), rat sections, marsupial PtK2 cells and hamster cell lines. The range of the number of dots and their size differed between cells and cell lines and varied between 1 and 24. The antigen could be identified as a 53 kDa protein with a pI of 8.7 and was named NDP53. Digestion experiments suggested that the protein is not associated with DNA or RNA, but is associated with the nuclear matrix. Immunelectron microscopy using ultra-thin sections revealed filamentous structures with a diameter of 0.1 to 0.6 microns. The antigen colocalizes with Sp100, PML and NDP55, which are part of a multiprotein complex known as PML oncogenic domain (POD), nuclear bodies, Kr bodies or ND10.

Animals↗

PubMed1996

Context dependent transactivation domains activate the immunoglobulin mu heavy chain gene enhancer.

Enhancers and promoters nucleate the assembly of multiprotein complexes that are required for the transcriptional activation of eukaryotic genes. Although multimerized binding sites of individual transcription factors sometimes mimic the properties of an enhancer, the combinatorial use of factors is considered to be crucial for achieving biological specificity. The minimal B cell specific immunoglobulin mu heavy chain gene enhancer is activated by a combination of tissue-restricted ETS proteins and ubiquitously expressed basic helix-loop-helix transcription factors. Here we show that a domain of PU.1 that activates transcription from multimerized PU.1 binding sites is not required to activate the mu enhancer together with Ets-1. In contrast, a transactivation domain in Ets-1 is necessary to activate this enhancer synergistically with PU.1. Furthermore, the Ets-1 activation domain functions only when tethered to the muA site of the enhancer. These observations illuminate two forms of context dependence: first, all possible transcription activation domains may not be required to achieve combinatorial specificity; second, functional transcription activation domains may require appropriate positioning on DNA.

Enhancer Elements, Genetic↗

PubMed1996

hTAF(II)68, a novel RNA/ssDNA-binding protein with homology to the pro-oncoproteins TLS/FUS and EWS is associated with both TFIID and RNA polymerase II.

TFIID is the main sequence-specific DNA-binding component of the RNA polymerase II (Pol II) transcriptional machinery. It is a multiprotein complex composed of the TATA-binding protein (TBP) and TBP-associated factors (TAF(II)s). Here we report the cloning and characterization of a novel human TBP-associated factor, hTAF(II)68. It contains a consensus RNA-binding domain (RNP-CS) and binds not only RNA, but also single stranded (ss) DNA. hTAF(II)68 shares extensive sequence similarity with TLS/FUS and EWS, two human nuclear RNA-binding pro-oncoproteins which are products of genes commonly translocated in human sarcomas. Like hTAF(II)68, TLS/FUS is also associated with a sub-population of TFIID complexes chromatographically separable from those containing hTAF(II)68. Therefore, these RNA and/or ssDNA-binding proteins may play specific roles during transcription initiation at distinct promoters. Moreover, we demonstrate that hTAF(II)68 co-purifies also with the human RNA polymerase II and can enter the preinitiation complex together with Pol II.

Amino Acid Sequence↗

PubMed1997

Cyclins and the G2/M transition.

The entry of a cell into mitosis is regulated by an elaborate network of kinases and phosphatases that control both for the timing of cell division and the complete reorganization of the cellular architecture. The mitotic cyclin/Cdks form part of large multiprotein complexes whose other components are only now beginning to be identified. The continuing identification of proteins that contribute to these complexes and changes in the composition of these complexes are likely to give a more integrated view of how mitotic cyclin/Cdk complexes are regulated and how they function-not only to induce mitosis, but also to aid further mitotic progression. Furthermore, assigning specific G2/M functions to distinct mitotic cyclin/Cdk complexes will require the identification of differences in substrate specificities between the mitotic cyclin/Cdk complexes, perhaps in parallel with specific cyclin knockouts in mice. Such investigations will be complicated by potential functional overlap between mitotic cyclin/Cdk complexes in vitro and in vivo. Although cyclin/Cdk1 is thought to be the major kinase that initiates the onset of mitosis, a more complete understanding of how cells move from G2 to a mitotic state will require further identification of kinases operating upstream, downstream and in parallel with Cdk1, their substrates and their relationship with one another during the G2/M transition.

Animals↗

PubMedDate not supplied

Role of hydroxyl radical in silica-induced NF-kappa B activation in macrophages.

Nuclear transcription factor kappa B (NF-kappa B) is a multiprotein complex that regulates a variety of genes important for immunity and inflammation. The present study investigates the silica-induced activation of this transcription factor in mouse macrophage cell line RAW 264.7 cells, the role of free radical reactions in the mechanism of the activation, and its possible inhibition. Tetrandrine, a benzylisoquinoline alkaloid, which has been used as an antifibrotic drug to treat the lesions of silicosis and has been characterized as a hydroxyl radical (.OH) scavenger, inhibited the NF-kappa B activation induced by silica, lipopolysaccharide (LPS), and phorbol 12-myristate 13-acetate (PMA). Catalase, metal chelator, deferoxamine, and the silanol group (SiOH) blocker, poly(2-vinylpyridine-N-oxide) (PVPNO), also inhibited silica-induced NF-kappa B activation. Electron spin resonance (ESR) spin trapping measurements show that both deferoxamine and PVPNO decreased silica-mediated .OH radical generation from H2O2. It is shown that Fe(II) and not Fe(III) is able to cause NF-kappa B activation. The antioxidant, ascorbate, attenuated the NF-kappa B activation induced by silica but not by LPS. The .OH radical scavenger, sodium formate, inhibited NF-kappa B activation induced by silica but had only a minor effect on NF-kappa B activation induced by LPS. The results indicate that silica-mediated free radical generation via the Fenton or Fenton-like reaction (M(n)+ + H2O2-->M(n + 1)+ + OH- + .OH) and silanol groups on the silica surface play an important role in silica-induced NF-kappa B activation.

Alkaloids↗

PubMed1997

Mapping specific protein-protein interactions within the core component of the breast cell DNA synthesome.

We have previously described the isolation and characterization of an intact multiprotein complex for DNA replication, designated the DNA synthesome, from human breast cancer cells and biopsied human breast tumor tissue. The purified DNA synthesome was observed to fully support DNA replication in vitro. We had also proposed a model for the breast cell DNA synthesome, in which DNA polymerases alpha, delta, and epsilon, DNA primase, and replication factor C (RF-C) represent members of the core component, or tightly associated, proteins of the complex. This model was based on the observed fractionation, chromatographic, and sedimentation profiles for these proteins. We report here that poly(ADP-ribose)polymerase (PARP) and DNA ligase 1 are also members of the breast cell DNA synthesome core component. More importantly, in this report we present the results of coimmunoprecipitation studies that were designed to map the protein-protein interactions between several members of the core component of the DNA synthesome. Consistent with our proposed model for the breast cell DNA synthesome, our data indicate that DNA polymerases alpha and delta, DNA primase, RF-C, as well as proliferating cell nuclear antigen (PCNA), tightly associate with each other in the complex, whereas DNA polymerase epsilon, PARP, and several other components were found to interact with the synthesome via a direct contact with only PCNA or DNA polymerase alpha. The association of PARP with the synthesome core suggests that this protein may serve a regulatory function in the complex. Also, the coimmunoprecipitation studies suggest that the three DNA polymerases alpha, delta, and epsilon all participate in the replication of breast cell DNA. To our knowledge this is the first report ever to describe the close physical association of polypeptides constituting the intact human breast cell DNA replication apparatus.

Breast↗

PubMed1998

UV light-induced cross-linking of nucleosides, nucleotides and a dinucleotide to the carboxy-terminal heptad repeat peptide of RNA polymerase II as studied by mass spectrometry.

We report here the results of a study to assess the usefulness of mass spectrometry as a method for rapidly locating cross-linking sites in peptides modified by UV irradiation in the presence of nucleic acid components. For this study, we selected two nucleosides (thymidine and 5-bromo-2'-deoxyuridine), two nucleotides (thymidine-5'-monophosphate and 5-bromo-2'-deoxyuridine-5'-monophosphate) and a dinucleotide (thymidylyl-[3'-->5']-2'-deoxyadenosine). The peptide picked was SPSYSPT (L-seryl-L-prolyl-L-seryl-L-tyrosyl-L-seryl-L-prolyl-L-threonine), the heptad repeat unit found in the largest subunit of the RNA polymerase II multiprotein complex. Modified peptides were isolated by reversed-phase HPLC. Molecular mass measurements confirmed that covalent adducts had been formed. High-energy tandem collision-induced dissociation mass spectrometry pinpointed the location of cross-linking in each modified peptide as being at the tyrosine residue. These results indicate that mass spectrometry is a potentially applicable technique for location of cross-linking sites in peptides, modified by attachment of nucleosides, nucleotides and dinucleotides. Such modified peptides would be among the products expected after application of standard proteolytic and nucleolytic digestion protocols to digestion of cross-linked DNA-protein complexes.

Amino Acid Sequence↗

PubMed1998

[Transcriptional control by steroid receptor].

Nuclear receptors for steroid/thyroid hormones, vitamin A,D and fat-soluble ligands form a gene superfamily of ligand-inducible transcription factors, which plays an important roles in a wide spectrum of biological events by regulating expression of a set of target genes. Two domains (A/B and E) of nuclear receptors are demonstrated to activate transcription, designated AF-1 (A/B) and AF-2 (E), respectively. Homo- and hetero-dimers of nuclear receptors bind target enhancer elements in the target gene promoters. The target enhancer elements are referred as hormone response elements (HRE), and composed of two hexamer core motifs. The orientations and spaces between two core motifs play critical roles in discreminating the recognition of HREs among the members of nuclear receptor. DNA-bound nuclear receptors controls transcription in a ligand-binding dependent way in co-operation with a multiprotein complex containing RNA polymerase II and a series of auxiliary factors, TFIIA,B,D, E,F and H. During process of ligand-induced transactivation by nuclear receptors, nuclear co-factors interacting with the AF-1 and AF-2 seem to be involved. Several transcriptional co-activators and co-repressors have been recently identified, and their function is currently under investigation.

Gene Expression↗

PubMedDate not supplied

DNMT1 is a component of a multiprotein DNA replication complex.

DNA methylation is a major determinant of epigenetic inheritance and plays an important role in genome stability. The accurate propagation of DNA methylation patterns with cell division requires that methylation be closely coupled to DNA replication, however the precise molecular determinants of this interaction have not been defined. In the present study, we show that the predominant DNA methyltransferase species in somatic cells, DNMT1, is a component of a multiprotein DNA replication complex termed the DNA synthesome that fully supports semi-conservative DNA replication in a cell-free system. DNMT1 protein and activity were found to co-purify with the human DNA synthesome through a series of subcellular fractionation and chromatography steps, resulting in an enrichment of methyltransferase specific activity from two human cell lines. DNA methyltransferase activity co-eluted with in vitro replication activity and DNA polymerase alpha activity on sucrose density gradients suggesting that DNMT1 is a tightly bound, core component of the replication complex. The synthesome-associated pool of DNA methyltransferase exhibited both maintenance and de novo methyltransferase activity and the ratio of the two was similar to that observed in whole cell lysates and for recombinant DNMT1. These data indicate that interactions within the synthesome complex do not influence the intrinsic preference of DNMT1 for hemimethylated DNA, but suggest that newly replicated DNA may be subject to low level de novo methylation. The data indicate that DNA methylation is tightly coupled to replication through physical interaction of DNMT1 and core components of the replication machinery. The definition of the molecular interactions between DNMT1 and other proteins in the replication complex in normal and neoplastic cells will provide further insight into the regulation of DNA methylation and the mechanisms underlying the alteration of DNA methylation patterns during carcinogenesis.

Cell Division↗

PubMed1996

The expression of poly(ADP-ribose) polymerase during differentiation-linked DNA replication reveals that it is a component of the multiprotein DNA replication complex.

3T3-L1 preadipocytes have been shown to exhibit a transient increase in poly(ADP-ribose) polymerase (PARP) protein and activity, as well as an association of PARP with DNA polymerase alpha, within 12-24 h of exposure to inducers of differentiation, whereas 3T3-L1 cells expressing PARP antisense RNA showed no increase in PARP and are unable to complete the round of DNA replication required for differentiation into adipocytes. The role of PARP in differentiation-linked DNA replication has now been further clarified at both the cellular and enzymological levels. Flow cytometric analysis revealed that control 3T3-L1 cells progressed through one round of DNA replication prior to the onset of terminal differentiation, whereas cells expressing PARP antisense RNA were blocked at the G0/G1 phase of the cell cycle. Confocal microscope image analysis of control S phase cells demonstrated that PARP was localized within distinct intranuclear granular foci associated with DNA replication centers. On the basis of these results, purified replicative complexes from other cell types that had been characterized for their ability to catalyze viral DNA replication in vitro were analyzed for the presence of PARP. PARP exclusively copurified through a series of centrifugation and chromatography steps with core proteins of an 18-21S multiprotein replication complex (MRC) from human HeLa cells, as well as with the corresponding mouse MRC from FM3A cells. The MRC were shown to contain DNA polymerases alpha and delta, DNA primase, DNA helicase, DNA ligase, and topoisomerases I and II, as well as accessory proteins such as PCNA, RF-C, and RP-A. Finally, immunoblot analysis of MRCs from both cell types with monoclonal antibodies to poly (ADP-ribose) revealed the presence of approximately 15 poly(ADP-ribosyl)ated proteins, some of which were further confirmed to be DNA polymerase alpha, DNA topoisomerase I, and PCNA by immunoprecipitation experiments. These results suggest that PARP may play a regulatory role within the replicative apparatus as a molecular nick sensor controlling the progression of the replication fork or modulates component replicative enzymes or factors in the complex by directly associating with them or by catalyzing their poly(ADP-ribosyl)ation.

3T3 Cells↗

PubMed1996

The human cell multiprotein DNA replication complex (MRC): the effect of camptothecin on its ability to support in vitro DNA synthesis.

PURPOSE: We have previously reported on the isolation and characterization of a multiprotein DNA replication complex (MRC) from HeLa cells that fully supports in vitro DNA replication. Based upon its ability to replicate DNA in a cell-free environment (devoid of other cellular processes) the MRC may serve as a unique model system for investigating the mechanisms of action of anticancer drugs that directly affect DNA synthesis. The experiments described in this report were performed to establish whether the MRC could serve as a model system to examine in detail the mechanism of action of camptothecin, a DNA topoisomerase I inhibitor. METHODS: We examined the effects of increasing concentrations of camptothecin on HeLa cell survival, intact HeLa cell DNA synthesis and MRC-mediated in vitro DNA replication. We also performed topoisomerase I assays in the presence of increasing concentrations of camptothecin to study the direct effects of the agent on MRC-associated topoisomerase I activity. Furthermore, we employed an SDS precipitation assay to measure the formation of MRC-associated topoisomerase I-cleavable complexes in the presence of increasing concentrations of camptothecin. RESULTS: We found a close correlation between the IC50 values for intact HeLa cell DNA synthesis (0.15 microM) and MRC-mediated in vitro DNA synthesis (0.05 microM). Similarly, we found that 0.05 microM camptothecin inhibited MRC-associated topoisomerase I activity by approximately 50%. In addition, we found that the formation of MRC-associated topoisomerase I-cleavable complexes increased linearly with increasing concentrations of camptothecin. CONCLUSIONS: The data presented in this report support the use of the MRC as a model system to study the mechanism of action of camptothecin. We anticipate that future studies with the MRC will help elucidate the cellular consequences of camptothecin-cleavable complex formation.

Antineoplastic Agents, Phytogenic↗

PubMed2004

Atomic force microscopy revelation of molecular complexes in the multiprotein cytochrome P450 2B4-containing system.

The application of atomic force microscopy (AFM) to the identification and visualization of individual molecules and their complexes in a reconstituted monooxygenase P450 2B4 system without the phospholipid was demonstrated. The method employed in this study distinguishes the monomeric proteins from their binary complexes and, also, the binary from the ternary complexes. The AFM images of the full-length P450 2B4 system's constituent components - cytochrome P450 2B4 (2B4), NADPH-cytochrome P450 reductase and cytochrome b5 (b5), were obtained on highly-oriented pyrolitic graphite. The typical heights of the d-2B4, d-flavoprotein (Fp) and d-b5 molecules were measured and found to be 2.2 +/- 0.2, 2.3 +/- 0.2 and 1.8 +/- 0.1 nm, respectively. The measured heights of the binary d-Fp/d-2B4 and d-2B4/d-b5 complexes were estimated to be 3.4 +/- 0.2 and 2.8 +/- 0.2 nm, respectively. No formation of d-Fp/d-b5 complexes was registered. The ternary d-Fp/d-2B4/d-b5 complexes were visualized and their heights were found to be roughly equal to 4.3 +/- 0.3 nm and 6.2 +/- 0.3 nm.

Animals↗

PubMed1996

Specific interaction of DNA polymerase beta and DNA ligase I in a multiprotein base excision repair complex from bovine testis.

Base excision repair (BER) is a cellular defense mechanism repairing modified bases in DNA. Recently, a G:U repair reaction has been reconstituted with several purified enzymes from Escherichia coli (Dianov, G., and Lindahl, T.(1994) Curr. Biol. 4, 1069-1076). Using bovine testis crude nuclear extract, we have shown that G:U is repaired efficiently in vitro, and DNA polymerase beta (beta-pol) is responsible for the single nucleotide gap-filling synthesis (Singhal, R. K., Prasad, R., and Wilson, S. H.(1995) J. Biol. Chem. 270, 949-957). To investigate potential interaction of beta-pol with other BER protein(s), we developed affinity chromatography matrices by cross-linking purified rat beta-pol or antibody against beta-pol to solid supports. Crude nuclear extract from bovine testis was applied to these affinity columns, which were then extensively washed. Proteins that bound specifically to the affinity columns were co-eluted in a complex with beta-pol. This complex had a molecular mass of approximately 180 kDa and was able to conduct the complete uracil-initiated BER reaction. The BER complex contained both beta-pol and DNA ligase I. An antibody to beta-pol was able to shift the complex in sucrose gradients to a much larger molecular mass (>300 kDa) that again contained both beta-pol and DNA ligase I. Furthermore, DNA ligase I and beta-pol were co-immunoprecipitated from the testis nuclear extract with anti beta-pol IgG. Thus, we conclude that beta-pol and DNA ligase I are components of a multiprotein complex that performs BER.

Animals↗