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At least 1,747 records · Page 97Linked to original sources
PubMed2005

Identification of a plastid acyl-acyl carrier protein synthetase in Arabidopsis and its role in the activation and elongation of exogenous fatty acids.

Plant cells are known to elongate exogenously provided fatty acid (FA), but the subcellular sites and mechanisms for this process are not currently understood. When Arabidopsis leaves were incubated with 14C-FAs with or=20 carbons) but not synthesis of 14C-unsaturated 18-carbon or 16-carbon FAs. Isolated pea chloroplasts were also able to elongate 14C-FAs (<or=16 carbons) in the light. No detectable 14C-acyl-CoA intermediates were formed during 14C-laurate elongation, whereas 14C-acyl-ACP accumulated to 2.3 microm. These data indicate that the elongation of exogenous medium-chain FAs to 16- and 18-carbon FAs occurs primarily in the chloroplasts, most likely via the enzymes of de novo FA synthesis. An Arabidopsis mutant with a T-DNA insertion in At4g14070 (AAE15) was reduced 80% in 14C-laurate elongation into 16- and 18-carbon FAs. AAE15 has sequence similarity to long-chain acyl-CoA synthetases and a predicted N-terminal plastidial targeting sequence. Direct acyl-ACP-forming activity from FA and ACP was observed in extracts of Arabidopsis leaves and isolated chloroplasts but aae15 plants had markedly reduced in vitro acyl-ACP synthesis activity. Together these results demonstrate that plants possess a mechanism for direct activation of FA to ACP in the plastid via an acyl-ACP synthetase encoded by At4g14070.

Arabidopsis↗
PubMed1999

Reciprocal interaction between two cellular proteins, Puralpha and YB-1, modulates transcriptional activity of JCVCY in glial cells.

Cross communication between regulatory proteins is an important event in the control of eukaryotic gene transcription. Here we have examined the structural and functional interaction between two cellular regulatory proteins, YB-1 and Puralpha, on the 23-bp sequence element derived from the enhancer-promoter of the human polyomavirus JCV. YB-1 and Puralpha are single-stranded DNA binding proteins which recognize C/T- and GC/GA-rich sequences, respectively. Results from band shift studies demonstrated that while both proteins interact directly with their DNA target sequences within the 23-bp motif, each protein can regulate the association of the other one with the DNA. Affinity chromatography and coimmunoprecipitation provide evidence for a direct interaction between Puralpha and YB-1 in the absence of the DNA sequence. Ectopic expression of YB-1 and Puralpha in glial cells synergistically stimulated viral promoter activity via the 23-bp sequence element. Results from mutational studies revealed that residues between amino acids 75 and 203 of YB-1 and between amino acids 85 and 215 of Puralpha are important for the interaction between these two proteins. Functional studies with glial cells indicated that the region within Puralpha which mediates its association with YB-1 and binding to the 23-bp sequence is important for the observed activation of the JCV promoter by the Puralpha and YB-1 proteins. The results of this study suggest that the cooperative interaction between YB-1 and Puralpha mediates the synergistic activation of the human polyomavirus JCV genome by these cellular proteins. The importance of these findings for cellular and viral genes which are regulated by Puralpha and YB-1 is discussed.

Binding Sites↗
PubMed2003

A second glutamine synthetase gene with expression in the gills of the gulf toadfish (Opsanus beta).

We characterized the expression of the nitrogen metabolism enzyme glutamine synthetase [GSase; L-glutamate: ammonia ligase (ADP-forming), E.C. 6.3.1.2] in tissues of the gulf toadfish Opsanus beta subjected to unconfined (ammonotelic) and confined (ureotelic) conditions. Enzymological results demonstrate that mass-specific GSase activities rank in the order of brain > liver > stomach approximately kidney > intestine > gill > heart/spleen > muscle. When tissue mass is used to calculate a glutamine synthetic potential, the liver has the greatest, followed by muscle > stomach and intestine, with minor contributions from the remaining tissues. Additionally, during confinement stress, GSase activity increases significantly only in liver (fivefold) and muscle (twofold), tissues that previously showed significant expression of the other enzymes of urea synthesis. Western analyses of samples on SDS gels demonstrated that GSase-specific protein content reflected enzyme activity, and all tissues except muscle had a single, similarly sized GSase subunit of 49.4 kDa; muscle showed staining of two bands of 36.8 and 98.9 kDa, which may possibly result from another gene product or post-translational modification. RT-PCR and RACE-PCR revealed the presence of a second GSase cDNA from gill tissue that shares only 73% nucleotide and amino acid sequence similarity with the GSase cDNA previously cloned from liver, and that lacks a mitochondrial leader-targeting sequence. RT-PCR and restriction digestion experiments demonstrated that mRNA from the original 'liver' GSase is expressed in all tissues examined (liver, gill, stomach, intestine, kidney, brain and muscle), whereas the new 'gill' form shows expression primarily in the gill. Gill GSase activity shows apparently exclusive expression in the soluble compartment, while other tissues expressing the 'liver' form show both cytoplasmic and mitochondrial activities. Phylogenetic analysis of a number of GSases demonstrates that the toadfish gill GSase has a greater affinity for a clade that includes the Xenopus GSase genes and one of two Fugu GSase genes, than it has for a clade containing the toadfish liver GSase and other described teleost GSase genes. The results are discussed in the context of a prior hypothesis on an ammonia-trapping mechanism in the gill of the toadfish.

Amino Acid Sequence↗
PubMed1994

Human mitochondrial transcription termination exhibits RNA polymerase independence and biased bipolarity in vitro.

Human mitochondrial 16 S rRNA 3'-end formation requires a tridecamer template sequence and a trans-acting protein of approximately 34 kDa. This protein binds tightly to its target sequence and further analysis of the protein-DNA complex revealed that the DNA is bent. Either T3, T7, Escherichia coli, or yeast mitochondrial RNA polymerase produced transcripts mapping at this termination site. With these heterologous RNA polymerase, RNA 3'-end formation was detected only in the transcription polarity opposite that of mitochondrial rRNA synthesis; the efficiency of termination in the homologous human RNA polymerase system is approximately 2-fold greater in this same opposite polarity. These results suggested the possible importance of biased bipolar transcription termination in vivo. For wild-type mtDNA, the apparent relative efficiency of termination in vivo reflected the values determined in vitro. Examination of a pathogenic human mtDNA mutation known to result in impaired termination in vitro showed no significant differences in relative transcript abundances in vivo, despite a loss of in vitro termination efficiency in both directions. Recently, six additional mitochondrial disease-associated point mutations have been reported that cluster at the human mitochondrial transcription termination site. None of these resulted in significantly impaired transcription termination in vitro.

Base Sequence↗
PubMed2004

Substrate twinning activates the signal recognition particle and its receptor.

Signal sequences target proteins for secretion from cells or for integration into cell membranes. As nascent proteins emerge from the ribosome, signal sequences are recognized by the signal recognition particle (SRP), which subsequently associates with its receptor (SR). In this complex, the SRP and SR stimulate each other's GTPase activity, and GTP hydrolysis ensures unidirectional targeting of cargo through a translocation pore in the membrane. To define the mechanism of reciprocal activation, we determined the 1.9 A structure of the complex formed between these two GTPases. The two partners form a quasi-two-fold symmetrical heterodimer. Biochemical analysis supports the importance of the extensive interaction surface. Complex formation aligns the two GTP molecules in a symmetrical, composite active site, and the 3'OH groups are essential for association, reciprocal activation and catalysis. This unique circle of twinned interactions is severed twice on hydrolysis, leading to complex dissociation after cargo delivery.

Amino Acid Sequence↗
PubMed2001

Crystal structure of the CCAAT box/enhancer-binding protein beta activating transcription factor-4 basic leucine zipper heterodimer in the absence of DNA.

The crystal structure of the heterodimer formed by the basic leucine zipper (bZIP) domains of activating transcription factor-4 (ATF4) and CCAAT box/enhancer-binding protein beta (C/EBP beta), from two different bZIP transcription factor families, has been determined and refined to 2.6 A. The structure shows that the heterodimer forms an asymmetric coiled-coil. Even in the absence of DNA, the basic region of ATF4 forms a continuous alpha-helix, but the basic region of C/EBP beta is disordered. Proteolysis, electrophoretic mobility shift assay, circular dichroism, and NMR analyses indicated that (i) the bZIP domain of ATF4 is a disordered monomer and forms a homodimer upon binding to the DNA target; (ii) the bZIP domain of ATF4 forms a heterodimer with the bZIP domain of C/EBP beta that binds the cAMP response element, but not CCAAT box DNA, with high affinity; and (iii) the basic region of ATF4 has a higher alpha-helical propensity than that of C/EBP beta. These results suggest that the degree of ordering of the basic region and the fork and the dimerization properties of the leucine zipper combine to distinguish the structurally similar bZIP domains of ATF4 and C/EBP beta with respect to DNA target sequence. This study provides insight into the mechanism by which dimeric bZIP transcription factors discriminate between closely related but distinct DNA targets.

Activating Transcription Factor 4↗
PubMed1993

RAMHA: a PC-based Monte-Carlo simulation of random saturation mutagenesis.

Random mutagenesis is a powerful tool in protein structure-function analyses. One approach to random mutagenesis is the de novo synthesis of polypeptide-encoding oligodeoxy-nucleotides using doped nucleoside phosphoramidites. A Turbo PASCAL program, RAMHA, is described for modeling such mutagenesis. Upon entering the target sequence and the desired level of nucleotide contamination, RAMHA performs a Monte Carlo simulation of the mutagenesis, compiling statistics on the similarity of resultant mutant polypeptides to the wild-type sequence, the frequency of premature open-reading frame terminations, and other relevant outcomes. Simulated mutagenesis of two DNA targets has led to the development of two different strategies to avoid the random introduction of stop codons within mutagenized gene segments.

Base Sequence↗
PubMed2006

CRZ1, a target of the calcineurin pathway in Candida albicans.

Calcineurin is a major player in calcium-dependent signal transduction pathways of eukaryotes. Calcineurin acts on transcription factors (e.g. CRZ1 in Saccharomyces cerevisiae) and governs the expression of genes in a species-dependent fashion. In Candida albicans, the calcineurin pathway is involved in tolerance to antifungal agents, cation homeostasis and virulence. However, the components of the calcineurin pathway are still poorly investigated in this yeast species. Taking S. cerevisiae as a model to reconstitute this pathway, two CRZ1-like genes, CRZ1 and CRZ2 (for calcineurin-responsive zinc finger 1 and 2 genes), were found with C(2)H(2) zinc finger domains. Only CRZ1 was able to restore the calcium hypersusceptibility of a S. cerevisiae crz1Delta mutant and to mediate calcium-dependent gene expression in this yeast species. Several experiments showed that CRZ1 was dependent on calcineurin in C. albicans: (i) phenotypic analysis of a crz1Delta/Delta mutant showed impaired growth as compared with the wild type in the presence of cations (Ca(2+), Mn(2+)) as does a mutant lacking calcineurin subunit A (cnaDelta/Delta) and (ii) a green fluorescent protein (GFP)-Crz1p fusion protein showed a calcium- and calcineurin-dependent nuclear localization. To further analyse the relationship between calcineurin and CRZ1, a comprehensive analysis of calcineurin/Crz1p-dependent gene expression following addition of Ca(2+) (200 mM) was performed. Among the expression of 264 genes altered by at least twofold, the upregulation of 60 genes was dependent on both calcineurin and CRZ1. Interestingly, a motif [5'-G(C/T)GGT-3'] with similarity to the target sequence of Crz1p (GNGGCG/TCA) from S. cerevisiae was identified as a putative regulatory sequence in the upstream regions of these calcineurin/Crz1p-dependent genes. However, additional experiments showed that calcineurin may have other targets in addition to CRZ1. First, CRZ1 was not involved in tolerance to antifungal agents (fluconazole, terbinafine) on the opposite to calcineurin. Second, CRZ1 was only moderately influencing virulence in a mice model of infection which is in sharp contrast to the strong avirulence of cnaDelta/Delta mutant in the same animal model. Even though this work establishes CRZ1 as a calcineurin target, further studies are needed to identify other calcineurin-dependent elements in C. albicans.

Animals↗
PubMed1985

Universal restriction endonucleases: designing novel cleavage specificities by combining adapter oligodeoxynucleotide and enzyme moieties.

Class IIS restriction endonucleases cleave double-stranded (ds) DNA at precise distances from their recognition sequences. A method is proposed which utilizes this separation between the recognition site and the cut site to allow a class IIS enzyme, e.g., FokI, to cleave practically any predetermined sequence by combining the enzyme with a properly designed oligodeoxynucleotide adapter. Such an adapter is constructed from the constant recognition site domain (a hairpin containing the ds sequence, e.g., GGATG CCTAC for FokI) and a variable, single-stranded (ss) domain complementary to the ss sequence to be cleaved (at 9 and 13 nucleotides on the paired strands from the recognition sequence in the example of FokI). The ss sequence designated to be cleaved could be provided by ss phage DNA (e.g., M13), gapped ds plasmids, or supercoiled ds plasmids that were alkali denatured and rapidly neutralized. Combination of all three components, namely the class IIS enzyme, the ss DNA target sequence, and the complementing adapter, would result in target DNA cleavage at the specific predetermined site. The target ss DNA could be converted to the precisely cleaved ds DNA by DNA polymerase, utilizing the adapter oligodeoxynucleotide as primer. This novel procedure represents the first example of changing enzyme specificity by synthetic design. A practically unlimited assortment of new restriction specificities could be produced. The method should have many specific and general applications when its numerous ramifications are exploited.

Base Sequence↗
PubMed1985

Enzymatic amplification of beta-globin genomic sequences and restriction site analysis for diagnosis of sickle cell anemia.

Two new methods were used to establish a rapid and highly sensitive prenatal diagnostic test for sickle cell anemia. The first involves the primer-mediated enzymatic amplification of specific beta-globin target sequences in genomic DNA, resulting in the exponential increase (220,000 times) of target DNA copies. In the second technique, the presence of the beta A and beta S alleles is determined by restriction endonuclease digestion of an end-labeled oligonucleotide probe hybridized in solution to the amplified beta-globin sequences. The beta-globin genotype can be determined in less than 1 day on samples containing significantly less than 1 microgram of genomic DNA.

Alleles↗
PubMed2001

Binding to the naturally occurring double p53 binding site of the Mdm2 promoter alleviates the requirement for p53 C-terminal activation.

Genotoxic stress activation of the tumor suppressor transcription factor p53 involves post-translational C-terminal modifications that increase both protein stability and DNA binding activity. We compared the requirement for p53 protein activation of p53 target sequences in two major p53-regulated genes, p21/WAF1 (encoding a cell cycle inhibitory protein) and Mdm2 (encoding a ubiquitin ligase that targets p53 for proteolytic degradation). The p53 binding site in the proximal p21/WAF1 promoter contains a single p53 binding consensus sequence, while the p53 binding site in the Mdm2 promoter contains two consensus sequences linked by a 17 bp spacer. Binding of recombinant p53 protein to the p21/WAF1 binding site required monoclonal antibody PAb421, which can mimic activating phosphorylation and/or acetylation events at the C-terminus. In contrast, recombinant p53 bound strongly to the Mdm2 binding site in the absence of PAb421 antibody. Separate binding to each consensus sequence of the Mdm2 binding site still required PAb421, indicating that p53 binding was not simply due to greater affinity to the Mdm2 consensus sequences. Linking two p21/WAF1 binding sites with the 17 bp spacer region from the Mdm2 gene eliminated the PAb421 requirement for p53 binding to the p21/WAF1 site. These results suggest a mechanism for regulation of Mdm2 gene transcription that differs from that other p53-induced genes by its lack of a requirement for C-terminal activation of p53 protein. A steady induction of Mdm2 protein would maintain p53 protein at low levels until post-translational modifications following DNA damage increased p53 activity towards other genes, mediating p53 growth inhibitory and apoptotic activities.

Base Sequence↗
PubMed1998

Crystal structure of the DpnM DNA adenine methyltransferase from the DpnII restriction system of streptococcus pneumoniae bound to S-adenosylmethionine.

BACKGROUND: . Methyltransferases (Mtases) catalyze the transfer of methyl groups from S-adenosylmethionine (AdoMet) to a variety of small molecular and macromolecular substrates. These enzymes contain a characteristic alpha/beta structural fold. Four groups of DNA Mtases have been defined and representative structures have been determined for three groups. DpnM is a DNA Mtase that acts on adenine N6 in the sequence GATC; the enzyme represents group alpha DNA Mtases, for which no structures are known. RESULTS: . The structure of DpnM in complex with AdoMet was determined at 1.80 A resolution. The protein comprises a consensus Mtase fold with a helical cluster insert. DpnM binds AdoMet in a similar manner to most other Mtases and the enzyme contains a hollow that can accommodate DNA. The helical cluster supports a shelf within the hollow that may recognize the target sequence. Modeling studies indicate a potential site for binding the target adenine, everted from the DNA helix. Comparison of the DpnM structure and sequences of group alpha DNA Mtases indicates that the group is a genetically related family. Structural comparisons show DpnM to be most similar to a small-molecule Mtase and then to macromolecular Mtases, although several dehydrogenases show greater similarity than one DNA Mtase. CONCLUSIONS: . DpnM, and by extension the DpnM family or group alpha Mtases, contains the consensus fold and AdoMet-binding motifs found in most Mtases. Structural considerations suggest that macromolecular Mtases evolved from small-molecule Mtases, with different groups of DNA Mtases evolving independently. Mtases may have evolved from dehydrogenases. Comparison of these enzymes indicates that in protein evolution, the structural fold is most highly conserved, then function and lastly sequence.

Amino Acid Sequence↗
PubMed2001

Sequence-specific delivery of a quinone methide intermediate to the major groove of DNA.

Silyl-protected phenol derivatives serve as convenient precursors for generating highly electrophilic quinone methide intermediates under biological conditions. Reaction is initiated by addition of fluoride and has previously exhibited proficiency in DNA alkylation and cross-linking. This approach has now been extended to the modification of duplex DNA through triplex recognition and fluoride-dependent quinone methide induction. Both oligonucleotides of a model duplex were alkylated in a sequence specific manner by an oligonucleotide conjugate that is consistent with triplex association. Optimum reaction required the presence of the two complementary target sequences and a pH of below 6.5. In addition, one guanine in each strand adjacent to the triplex region was the predominant site of alkylation. The yield of modification varied from approximately 20% for the purine-rich strand to only 4% for the pyrimidine-rich strand. This surprising difference indicates that the linker between the recognition and reactive elements may limit productive interaction between the quinone methide and the reactive nucleophiles of DNA. Restricted orientation of this intermediate may also be responsible for the lack of target cross-linking at detectable levels.

Alkylation↗
PubMed1996

A Rho-dependent transcription termination site regulated by bacteriophage P4 RNA immunity factor.

The genes required for replication of the temperate bacteriophage P4, which are coded by the phage left operon, are expressed from a constitutive promoter (PLE). In the lysogenic state, repression of the P4 replication genes is achieved by premature transcription termination. The leader region of the left operon encodes all the genetic determinants required for prophage immunity, namely: (i) the P4 immunity factor, a short, stable RNA (CI RNA) that is generated by processing of the leader transcript; (ii) two specific target sequences that exhibit complementarity with the CI RNA. RNA-RNA interactions between the CI RNA and the target sites on the mRNA leader region are essential for transcription termination. To understand how transcription termination is elicited by the P4 immunity mechanism, it is relevant to identify the transcription termination site. This, however, could not be directly inferred from the 3'-end of the transcription products because of the extensive and complex processing and degradation of the leader RNA. In this work, by making use of a tRNA gene as a reporter, we identify the termination site of the immunity transcripts (timm). This is a Rho-dependent terminator located within the first translated gene of the left operon and is regulated by P4 immunity. Analysis of the P4 transcription pattern in Escherichia coli rho mutants suggests that termination at timm may also be important for the efficient processing of the CI RNA.

Base Sequence↗
PubMed1994

Roles of transcription and repair in alkylation mutagenesis.

Mutations occurring in Escherichia coli cells exposed to alkylating agents have been analyzed using an assay for forward mutations in the E. coli rpsL gene cloned on a high copy number plasmid. N-Methyl-N'-nitro-N-nitrosoguanidine (MNNG)-induced mutations were recovered from wild-type and O6-methylguanine methyltransferase-deficient mutant (ada- ogt-) cells and their sequence alterations determined. We found that the mutations recovered from the wild-type strain were predominantly G:C to A:T transitions located at several hot spots in the rpsL sequence. A vast majority of the mutations were found at guanine residues preceded by thymine on the transcribed strand of the target gene. Although the methyltransferase mutant showed hypersensitivity to the alkylating reagent in terms of mutagenic effect and cell killing effects, the class and site distributions of the rpsL- mutations recovered from MNNG-treated ada- ogt- cells were similar to those observed with MNNG-treated wild-type cells. Therefore, the site preference of MNNG-induced rpsL- mutations seems to be due not to the specificity of methyl-transferring repair enzymes but probably to the distribution of the mutagenic lesions (O6-methylguanine) in the target sequence. Mutations induced by methyl methanesulfonate, an SN2 alkylating agent, showed similar class and site distributions in the rpsL system. The site preference of MNNG-induced mutations was significantly changed when the level of transcription of the rpsL gene was decreased to 120-fold lower than that promoted by the authentic rpsL promoter. Under these conditions, 78% of mutations were induced at the central guanine of 5'-GG(A or C)-3' and 2/3 of them were on the non-transcribed strand of the rpsL gene. These results suggested that the site preference of MNNG-induced mutations is determined by at least three factors: (i) a flanking-base effect on the chemical reactivity of a guanine residue, (ii) transcribed strand-specific repair, probably by the UvrABC system, and (iii) the effects of transcription of the target gene on the alkylation of DNA and the strand-specific repair.

Alkylation↗
PubMed2005

Complex protein targeting to dinoflagellate plastids.

Protein trafficking pathways to plastids are directed by N-terminal targeting peptides. In plants this consists of a relatively simple transit peptide, while in organisms with secondary plastids (which reside within the endomembrane system) a signal peptide is appended to the transit peptide. Despite amino acid compositional differences between organisms, often due to nucleotide biases, the features of plastid targeting sequences are generally consistent within species. Dinoflagellate algae deviate from this trend. We have conducted an expressed sequence tag (EST) survey of the peridinin-plastid containing dinoflagellate Heterocapsa triquetra to identify and characterize numerous targeting presequences of plastid proteins encoded in the nucleus. Consistent with targeting systems present in other secondary plastid-containing organisms, these all possess a canonical signal peptide at their N termini, however two major classes of transit peptides occur. Both classes possess a common N-terminal portion of the transit peptide, but one class of transit peptides contains a hydrophobic domain that has been reported to act as a stop-transfer membrane anchor, temporarily arresting protein insertion into the endoplasmic reticulum. A second class of transit peptide lacks this feature. These two classes are represented approximately equally, and for any given protein the class is conserved across all dinoflagellate taxa surveyed to date. This dichotomy suggests that two mechanisms, perhaps even trafficking routes, may direct proteins to dinoflagellate plastids. A four-residue phenylalanine-based motif is also a consistent feature of H. triquetra transit peptides, which is an ancient feature predating red algae and galucophytes that was lost in green plastids.

Amino Acid Sequence↗
PubMed1998

Binding of the fur (ferric uptake regulator) repressor of Escherichia coli to arrays of the GATAAT sequence.

The mode of DNA binding of the Fur (ferric uptake regulator) repressor which controls transcription of iron-responsive genes in Escherichia coli, has been re-examined. Using as a reference the known sites at the promoter of the aerobactin operon of Escherichia coli, we have compared in detail the patterns of interaction between the purified Fur protein and natural or synthetic DNA targets. DNase I and hydroxyl radical footprinting, as well as missing-T assays, consistently revealed that functional Fur sites are composed of a minimum of three repeats of the hexameric motif GATAAT rather than by a palindromic 19 bp target sequence. Extended binding sites, constructed by stepwise addition of one or two direct repeats of the same sequence, were occupied co-operatively by Fur with the same pattern of interactions as those observed with the core of three repeats. This indicated that functional sites with a range of affinities can be formed by the addition of discrete GATAAT extensions to a minimal recognition sequence. The fashion in which Fur binds its target, virtually unknown in prokaryotic transcriptional regulators, accounts for the observed helical wrapping of the protein around the DNA helix.

Bacterial Proteins↗
PubMed1990

Human immunodeficiency virus integration in a cell-free system.

Integration of the viral genome into the nuclear DNA of a host cell plays a pivotal role in the replication of retroviruses. We have developed an in vitro method for studying the biochemistry of human immunodeficiency virus (HIV) integration by using extracts from HIV-infected cells. Analysis of the reaction products showed that HIV integration in vitro accurately reproduces the in vivo process. Integration occurred without apparent specificity for the target sequence, and the integrated provirus was directly flanked by a 5-base-pair duplication of DNA from the target site. HIV integration did not require a high-energy cofactor, and the enzymatic activities required for integration were recovered with the viral DNA when cell extracts were fractionated by gel exclusion chromatography.

Base Sequence↗