Publications

hu and integration host factor function as auxiliary proteins in cleavage of phage lambda cohesive ends by terminase.	hu and integration host factor (ihf) are small, basic heterodimeric dna-binding proteins which participate in transcription initiation, dna replication, and recombination. we constructed isogenic escherichia coli strains in which hu, ihf, or both proteins were absent. bacteriophage lambda did not grow in hosts lacking both hu and ihf. phage dna replication and late gene transcription were normal in the double mutants, but packaging of lambda dna was defective. mature phage dna molecules were abs ...	1991	1825651
isolation and preliminary characterization of escherichia coli mutants resistant to lethal action of the bacteriophage lambda p gene.	both spontaneous and ntg-induced mutants of escherichia coli 594 insensitive to the lethal action of lambda p gene were isolated and called rpl (resistant to p lethality). these mutants were of two types, showing different phenotypes. on type i rpl mutants, lambda cl- and lambda v1v3 did not plate, while lambda vir, lambda cl- c17, lambda imm434, and lambda imm21 did; plasmid pmr45 carrying the lambda p gene could not complement lambda imm21p- phage in type i mutants. on the other hand, the type ...	1991	1827225
identification of protein binding sites in genomic dna by two-dimensional gel electrophoresis.	we describe a simple two-dimensional electrophoresis procedure to identify the recognition sites of dna-binding proteins within large dna molecules. using this approach, we have mapped e. coli ihf (integration host factor) binding sites within phage lambda (48 kb) and phage mu (39 kb) dna. we are also able to visualize ihf binding sites in e. coli chromosomal dna (4,700 kb). we present an extension of this technique using direct amplification by pcr of the isolated restriction fragments, which s ...	1991	1827523
heat-inducible reactivation of uv-damaged bacteriophage lambda.	induction of the sos response in uv-irradiated bacteria leads to an increase in the survival of an infecting irradiated bacteriophage lambda (weigle 1953). we report that a similar reactivation of irradiated phage lambda was induced by shifting the culture of recipient bacteria from 30 degrees to 47 degrees c. however, this repair process was nonmutagenic. the amplitude of the phenomenon was increased with the quantity of uv lesions in the phage dna. it was present despite mutations affecting th ...	1991	1827875
dna recognition by the helix-turn-helix motif: investigation by laser raman spectroscopy of the phage lambda repressor and its interaction with operator sites ol1 and or3.	the lambda repressor provides a model system for biophysical studies of dna recognition by the helix-turn-helix motif. we describe laser raman studies of the lambda operator sites ol1 and or3 and their interaction with the dna-binding domain of lambda repressor (residues 1-102). raman spectra of the two dna sites exhibit significant differences attributable to interstrand purine-purine steps that differ in the two oligonucleotides. remarkably, the conformation of each operator is significantly a ...	1991	1828373
heteroduplex chain polarity in recombination of phage lambda by the red, recbcd, recbc(d-) and recf pathways.	we have examined the chain polarity of heteroduplex dna in unreplicated, bacteriophage lambda splice recombinants when recombination was by the recbcd, recbc(d-), or recf pathway of escherichia coli or the red pathway of lambda. for each of these pathways, recombination is activated by the cutting of cos that accompanies chromosome packaging, and is effected by recombination enzymes acting at the right end created by that cutting. for exchanges occurring near cos, one parent makes a lesser physi ...	1991	1829428
efficient excision of phage lambda from the escherichia coli chromosome requires the fis protein.	the escherichia coli protein fis has been shown to bind a single site in the recombination region of phage lambda and to stimulate excisive recombination in vitro (j. f. thompson, l. moitoso de vargas, c. koch, r. kahmann, and a. landy, cell 50:901-908, 1987). we demonstrate that mutant strains deficient in fis expression show dramatically reduced rates of lambda excision in vivo. phage yields after induction of a stable lysogen are reduced more than 200-fold in fis cells. the defect observed in ...	1991	1829453
the last duplex base-pair of the phage lambda chromosome. involvement in packaging, ejection and routing of lambda dna.	cosn is the site at which the bacteriophage lambda dna packaging enzyme, terminase, introduces staggered nicks to generate the cohesive ends of mature lambda chromosomes. genetic and molecular studies show that cosn is recognized specifically by terminase and that effects of cosn mutations on lambda dna packaging and cosn cleavage are well correlated. mutations affecting a particular base-pair of cosn are unusual in being lethal in spite of causing only a moderate defect in cosn cleavage and dna ...	1991	1830344
isolation and characterization of mutations in the bacteriophage lambda terminase genes.	the terminase enzyme of bacteriophage lambda is a hetero-oligomeric protein which catalyzes the site-specific endonucleolytic cleavage of lambda dna and its packaging into phage proheads; it is composed of the products of the lambda nul and a genes. we have developed a simple method to select mutations in the terminase genes carried on a high-copy-number plasmid, based on the ability of wild-type terminase to kill reca strains of escherichia coli. sixty-three different spontaneous mutations and ...	1991	1830578
molecular cloning and nucleotide sequence of the rhizobium phaseoli reca gene.	a recombinant lambda phage carrying the reca gene of rhizobium phaseoli was isolated from a r. phaseoli genomic library by complementation of the fec- phenotype of the recombinant phage in escherichia coli. when expressed in e. coli, the cloned reca gene was shown to restore resistance to both uv irradiation and the dna alkylating agent methyl methanesulphonate (mms). the r. phaseoli reca gene also promoted homologous recombination in e. coli. the cloned reca gene was only weakly inducible in e. ...	1991	1832737
dna sequence changes in mutations induced by ultraviolet light in the gpt gene on the chromosome of escherichia coli uvr+ and urva cells.	sequence changes in mutations induced by ultraviolet light are reported for the chromosomal escherichia coli gpt gene in almost isogenic e. coli uvr+ and excision-deficient uvra cells. differences between the mutagenic spectra are ascribed to preferential removal of photoproducts in the transcribed strand by excision repair in uvr+ cells. this conclusion is confirmed by analysis of published results for genes in both uvr+ and uvr- cells, showing a similar selective removal of mutagenic products ...	1991	1836051
[synthesis, secretion, and proteolytic degradation of diphtheria toxin in escherichia coli].	the recombinant plasmids have been constructed encoding the synthesis of a full-sized diphtheria toxin from its own or pr, pl-promoters of bacteriophage lambda in escherichia coli cells. the high level constitutive synthesis of toxin results in slow cell growth and plasmid elimination. the toxin was mainly detected in the periplasm, partially in the membrane and to a less extent in the cytoplasm and culturing medium. the dimeric form of toxin was found in the cytoplasm. participation of toxin b- ...	1991	1836836
the kila operon of promiscuous plasmid rk2: the use of a transducing phage (lambda pklaa-1) to determine the effects of the lethal klaa gene on escherichia coli cells.	the kil-kor regulon of promiscuous plasmid rk2 includes the replication initiator gene trfa and several potentially host-lethal kil loci (kila, kilb, kilc, kile), whose functions may be involved in plasmid maintenance or broad host range. the kila locus consists of a single operon of three genes (klaa, klab, klac), each of which is lethal when expressed from the klaa promoter in the absence of repressors encoded by kora and korb. in this study, we examined the effects of the unregulated klaa gen ...	1991	1838127
recombinagenic processing of uv-light photoproducts in nonreplicating phage dna by the escherichia coli methyl-directed mismatch repair system.	nonreplicating lambda phage dna in homoimmune escherichia coli lysogens provides a useful model system for study of processes that activate dna for homologous recombination. we measured recombination by extracting phage dna from infected cells, using it to transfect reca recipient cells, and scoring the frequency of recombinant infective centers. with unirradiated phage, recombinant frequencies were less than 0.1%. however, recombination could be increased over 300-fold by prior uv irradiation o ...	1991	1838344
selection of lacz operon fusions in genes of gluconate metabolism in e. coli. characterization of a gntt::lacz fusion.	the initial steps involved in the utilization of gluconate by e. coli, its incorporation into the cell and subsequent phosphorylation to gluconate 6-phosphate, conform two systems that duplicate activities. these systems, gnti and gntii, are specified by two sets of genes distinctly regulated and located respectively at the mala-asd (75 min) and fdp-vals (96 min) regions of the bacterial chromosome. the presence of duplicate activities in the metabolism of gluconate of e. coli, has made difficul ...	1991	1843569
bacteriophage lambda promoters pl and pr: sequence determinants of in vivo activity and of sensitivity to the dna gyrase inhibitor, coumermycin.	sequence encompassing the region between bp -43 and +8 of the pl and pr promoters of bacteriophage lambda, as well as sequence variants of these promoters, were compared with respect to their ability to drive a promoterless cat gene in vivo. for both pl- and pr-based promoters, variants with one nonconsensus bp rather than the consensus promoter were found to be maximally active. determination of promoter function in csh26 and c600 revealed a marked strain dependence in the activity of some prom ...	1991	1847348
rna polymerases from pseudomonas aeruginosa and pseudomonas syringae respond to escherichia coli activator proteins.	the activities of rna polymerases (rnaps) from pseudomonas aeruginosa and pseudomonas syringae were compared with that of escherichia coli rnap. all three enzymes are able to initiate transcription at the trpba promoter of p. aeruginosa and at the coliphage lambda promoters, prm and pre, in response to heterospecific activators (trpi protein, repressor, and cii protein, respectively). however, both pseudomonas polymerases have less stringent requirements for promoter recognition in the absence o ...	1991	1898924
cloning, sequencing and overexpression of the gene for prokaryotic factor ef-p involved in peptide bond synthesis.	a soluble protein ef-p (elongation factor p) from escherichia coli has been purified and shown to stimulate efficient translation and peptide-bond synthesis on native or reconstituted 70s ribosomes in vitro. based on the partial amino acid sequence of ef-p, 18- and 24-nucleotide dna probes were synthesized and used to screen lambda phage clones from the kohara gene bank. the entire ef-p gene was detected on lambda clone #650 which contains sequences from the 94 minute region of the e.coli genome ...	1991	1956781
renaturation of cobra venom phospholipase a2 expressed from a synthetic gene in escherichia coli.	cobra venom (naja naja naja) phospholipase a2 (pla2) contains 14 cysteines in the form of 7 disulfide bonds amongst its 119 amino acids. a gene encoding the pla2 was synthesized and inserted into a bacterial expression vector containing the phage lambda pl promoter. in order to obtain protein without the initiating methionine at the n-terminus, a factor xa site was engineered upstream from the pla2 gene. upon heat-induction of the cells transformed with the expression plasmid, the protein is pro ...	1992	1730025
saccharomyces cerevisiae elongation factor 2. genetic cloning, characterization of expression, and g-domain modeling.	the elongation factor 2 (ef-2) genes of the yeast saccharomyces cerevisiae have been cloned and characterized with the ultimate goal of gaining a better understanding of the mechanism and control of protein synthesis. two genes (eft1 and eft2) were isolated by screening a bacteriophage lambda yeast genomic dna library with an oligonucleotide probe complementary to the domain of ef-2 that contains diphthamide, the unique posttranslationally modified histidine that is specifically adp-ribosylated ...	1992	1730643
spontaneous transposition in the bacteriophage lambda cro gene residing on a plasmid.	a new mutagenesis assay system based on the phage lambda cro repressor gene residing on a plasmid was developed. the assay detects mutations in cro that decrease the binding of the repressor to the or operator in an or pr-lacz fusion present in a lambda prophage. mutations arose spontaneously during growth of e. coli cells harboring cro plasmids at a frequency of 3-6 x 10(-6). analysis of some 200 cro mutants from several 'wild-type' strains revealed a substantial fraction of 25-70% insertion ev ...	1992	1373849
both forms of translational initiation factor if2 (alpha and beta) are required for maximal growth of escherichia coli. evidence for two translational initiation codons for if2 beta.	the gene infb codes for two forms of translational initiation factor if2; if2 alpha (97,300 da) and if2 beta (79,700 da). if2 beta arises from an independent translational event on a gug codon located 471 bases downstream from if2 alpha start codon. by site-directed mutagenesis we constructed six different mutations of this gug codon. in all cases, if2 beta synthesis was variably affected by the mutations but not abolished. we show that the residual expression of if2 beta results from translatio ...	1992	1374802
characterization of the transcription activator protein c1 of bacteriophage p22.	we cloned, expressed, and purified the positive regulatory protein c1 of the temperate phage p22 of salmonella typhimurium. the purified protein was characterized as to its amino acid composition, protein sequence, molecular weight, and antigenicity. p22 c1 was shown to be a tetrameric protein composed of four identical subunits with m(r) = 10,000. moreover, we identified and characterized two p22 c1-dependent phage promoters, p(re) and pa23, whose function was completely dependent on c1 both in ...	1992	1385814
construction of coliphage lambda charon vectors with bamh1 cloning sites. 1980.		1992	1422018
overproduction and purification of bacillus subtilis dna polymerase iii.	the objectives of this work were to engineer the cloned polc gene encoding bacillus subtilis dna polymerase iii for controlled overexpression in escherichia coli and to devise a facile purification scheme permitting the large-scale production of pure recombinant polymerase. the translational signals of polc were restructured by expression cassette pcr (macferrin et al., 1990, proc. natl. acad. sci. usa 87, 1937-1941), and the modified gene was inserted into the expression plasmid, pkc30 (rosenbe ...	1992	1422209
the translation initiation site of recombinant trypanosoma brucei ornithine decarboxylase varies with different promoters.	expression of the trypanosoma brucei ornithine decarboxylase (odc) gene in escherichia coli behind the lambda phage pr promoter led to the production of a recombinant enzyme having the same subunit molecular weight as the native enzyme [4]. however, when the same gene is expressed behind the tac promoter or the phoa promoter, the odcs produced by the transformed e. coli have subunit molecular weights approximately 2 kda higher than that of the native enzyme. amino terminal sequencing of the reco ...	1992	1435879
bacteriophage lambda papa: not the mother of all lambda phages.	the common laboratory strain of bacteriophage lambda--lambda wild type or lambda papa--carries a frameshift mutation relative to ur-lambda, the original isolate. the ur-lambda virions have thin, jointed tail fibers that are absent from lambda wild type. two novel proteins of ur-lambda constitute the fibers: the product of stf, the gene that is disrupted in lambda wild type by the frameshift mutation, and the product of gene tfa, a protein that is implicated in facilitating tail fiber assembly. r ...	1992	1439823
involvement of the escherichia coli rna polymerase alpha subunit in transcriptional activation by the bacteriophage lambda ci and cii proteins.	escherichia coli cells harbouring the rpoa341 mutation produce an rna polymerase which transcribes inefficiently certain operons subject to positive control. here, we demonstrate that the rpoa341 allele also prevents lysogenization of the host strain by bacteriophage lambda, a process dependent upon the action of two phage-encoded activators. this phenomenon was shown to arise from an inability to establish an integrated prophage rather than a failure to maintain the lysogenic state. the inabili ...	1992	1452017
enriched sources of escherichia coli replication proteins. the dnag primase is a zinc metalloprotein.	primase, the product of the escherichia coli dnag gene, is the enzyme responsible for rna primer synthesis on both template strands at replication forks during chromosomal dna synthesis. the dnag gene was modified by replacement of the natural ribosome-binding site with one complementary to the 3' end of 16s rrna, and then inserted downstream of tandem bacteriophage lambda pr and pl promoters in the puc9-derived vector pce30. following thermal induction of transcription, the resulting plasmid pp ...	1992	1511009
effect of escherichia coli nusg function on lambda n-mediated transcription antitermination.	the escherichia coli nus factors act in conjunction with the bacteriophage lambda n protein to suppress transcription termination on the lambda chromosome. nusa binds both n and rna polymerase and may also interact with other nus factors. to search for additional components of the n antitermination system, we isolated host revertants that restored n activity in nusa1 mutants. one revertant, nusg4, was mapped to the rif region of the e. coli chromosome and shown to represent a point mutation near ...	1992	1531224
lambda int protein bridges between higher order complexes at two distant chromosomal loci attl and attr.	the excisive recombination reaction of bacteriophage lambda involves a specific and efficient juxtaposition of two distant higher order protein-dna complexes on the chromosome of escherichia coli. these complexes, which mediate synapsis and strand exchange, consist of two dna sequences, attl and attr, the bivalent dna binding protein int, and the sequence-specific dna bending proteins, ihf, xis, and fis. the protein-protein and protein-dna interactions within, and between, these complexes were s ...	1992	1533056
an efficient phage plaque screen for the random mutational analysis of the interaction of hiv-1 gp120 with human cd4.	a lambda phage expression methodology was adapted to dissect protein/ligand interactions efficiently through the creation and rapid screening of large numbers of mutants. here we describe the method and its specific application to the interaction between the external envelope glycoprotein of the human immunodeficiency virus (hiv-1), gp120, and the human cell surface protein cd4. random substitutions were introduced throughout the gp120 binding region (amino acids 38-62) in the amino-terminal dom ...	1992	1533631
t41 mutation in lac repressor is tyr282----asp.	a monomeric mutant of the lac repressor protein, designated t41, produced originally by the in vivo mutt mutagenesis exhibited behavior similar to mutants identified as tyr282----ser or tyr282----gln [schmitz et al., j. biol. chem. 251 (1976) 3359-3366]. the t41 gene, encoded within a phage lambda prophage in the escherichia coli genome, was amplified by polymerase chain reaction and sequenced. the only mutation found in the nucleotide sequence corresponded to position 282 in the protein, and th ...	1992	1547952
cloning and nucleotide sequence of the escherichia coli cytidine deaminase (ccd) gene.	the structural gene that encodes cytidine deaminase (cdd) in escherichia coli was cloned from kohara phage lambda 365 (7f1), and its nucleotide sequence was determined. plasmids harboring the gene complemented chromosomal cdd mutations, enhanced cytidine deaminase activity in cell extracts, and directed the synthesis of a protein identical in mass and n-terminal amino acid sequence with cytidine deaminase purified from wild-type bacteria. metal analysis of the purified, plasmid-encoded deaminase ...	1992	1567863
cloning and overexpression of the lactobacillus bulgaricus nad(+)-dependent d-lactate dehydrogenase gene in escherichia coli:purification and characterization of the recombinant enzyme.	the lactobacillus bulgaricus nad(+)-dependent d-lactate dehydrogenase gene was amplified by the polymerase chain reaction and cloned into an escherichia coli expression plasmid pkk223.3. attempts to clone the full-length chromosomal dna encoding d-lactate dehydrogenase from a partial sau3ai lambda phage library or an enriched clone bank in e. coli were unsuccessful. the recombinant plasmid pkbuldh containing the amplified gene overexpressed d-lactate dehydrogenase (greater than 30% of total solu ...	1992	1610363
phage lambda has an analog of escherichia coli reco, recr and recf genes.	the recf pathway catalyzes generalized recombination in escherichia coli that is mutant for recbc, sbcb and sbcc. this pathway operating on conjugational recombination requires the reca, recf, recj, recn, reco, recq, recr, ruva, ruvb and ruvc genes. in contrast, lambda mutant for its own recombination genes, int, red alpha and red beta, requires only the reca and recj genes to recombine efficiently in recbc sbcb sbcc cells. deletion of an open reading frame in the ninr region of lambda results i ...	1992	1310087
alleviation of ecok dna restriction in escherichia coli and involvement of umudc activity.	the activity of the ecok dna restriction system of escherichia coli reduces both the plating efficiency of unmodified phage lambda and the transforming ability of unmodified pbr322 plasmid dna. however, restriction can be alleviated in wild-type cells, by uv irradiation and expression of the sos response, so that 10(3)- to 10(4)-fold increases in phage growth and fourfold increases in plasmid transformation occurred with unmodified dna. restriction alleviation was found to be a transient effect ...	1992	1310522
nonrandom orientation of transposon tn5supf insertions in phage lambda.	transposition of mini-transposon tn5supf to phage lambda can be selected in two ways: (i) by plaque formation on a dnab amber strain of escherichia coli, which requires expression of the transposon-borne suppressor trna gene (supf) during lytic phage growth, or (ii) by lysogenization of a strain with amber mutations in tet and amp resistance genes, and selection of tcr apr (sup+) transductant colonies. tn5supf insertions in several lambda clones were isolated and mapped using a polymerase chain ...	1992	1316868
the construction of streptomyces cyaneus genomic libraries in escherichia coli is dependent upon the use of mcr-deficient strains.	streptomyces cyaneus genomic dna ligated into either lambda phage or plasmid vectors was very inefficiently cloned into standard escherichia coli host strains. however, the same material could be efficiently cloned using mcr-deficient e. coli strains. these results suggest that the s. cyaneus genome contains 5-methylcytosine residues, some of which occur within the recognition sequences of the e. coli mcr restriction system.	1992	1327960
a selective lambda phage cloning vector with automatic excision of the insert in a plasmid.	a bacteriophage lambda cloning vehicle has been constructed for the generation of cdna libraries. the vector has the following properties. (1) it has a unique bamhi site engineered into the lambda gam gene. segments of dna can be cloned into this site and clones with an insert can be selected by their ability to grow on an escherichia coli host lysogenic for phage p2 (spi- phenotype). (2) when the recombinant phage infects a cre-producing e. coli strain, a site-specific recombination event resul ...	1992	1327972
efficient large-scale sequencing of the escherichia coli genome: implementation of a transposon- and pcr-based strategy for the analysis of ordered lambda phage clones.	we have developed a strategy for efficient sequence analysis of the genome of e. coli k-12 using insertions of a tn5-derived mini-transposon into overlapping ordered lambda phage clones to provide universal primer-binding sites, and pcr amplification of dna segments adjacent to the insertions. transposon-containing clones were selected by blue plaque formation on a dnabamber laczamber e. coli strain. insertion points every 0.5-1 kb were identified by 'analytical pcr' and segments between the tra ...	1992	1336178
characterization of gene expression in recombinant escherichia coli cells infected with phage lambda.	phage lambda infection was investigated for possible production of toxic foreign proteins in escherichia coli. the target gene transcription was regulated by a t7 promoter, which was initiated under the action of t7 rna polymerase delivered by infecting phage. two types of phage infection were investigated. in both cases, deletion of the int gene prevents lysogenic integration. one phage, lambda ce6, contains the sam7 lysis mutation, so that infectious phage particles remain intracellular. the o ...	1993	7763591
recognition of boxa antiterminator rna by the e. coli antitermination factors nusb and ribosomal protein s10.	the boxa sequences of the e. coli ribosomal rna (rrn) operons are sufficient to cause rna polymerase to read through rho-dependent transcriptional terminators. we show that a complex of the transcription antitermination factors nusb and ribosomal protein s10 interacts specifically with boxa rna. neither nusb nor s10 binds boxa rna on its own, and neither nusa nor nusg affects the interaction of the nusb-s10 complex with boxa rna. mutations in boxa that impair its antitermination activity comprom ...	1993	7678781
in vivo regulatory responses of four escherichia coli operons which encode leucyl-trnas.	four escherichia coli operons, the leuv operon which encodes trna(1leu), the leux operon which encodes trna(6leu), the mett operon which encodes trna(3leu), and the argt operon which encodes trna(1leu), were examined for the stringent response induced by serine hydroxamate and for growth rate-dependent regulation. in nuclease protection assays, the leuv operon displayed the stringent response in response to leucine starvation, analog inhibition, and growth of a temperature-sensitive leucyl-trna ...	1993	7680341
targeted mutagenesis of dna using triple helix-forming oligonucleotides linked to psoralen.	oligonucleotides can bind as third strands of dna in a sequence-specific manner in the major groove in homopurine/homopyrimidine stretches in duplex dna. here we use a 10-base triplex-forming oligonucleotide linked to a psoralen derivative at its 5' end to achieve site-specific, targeted mutagenesis in an intact, double-stranded lambda phage genome. site-specific triplex formation delivers the psoralen to the targeted site in the lambda dna, and photoactivation of the psoralen produces adducts a ...	1993	8356097
localization and nucleotide sequences of genes mediating site-specific recombination of the slp1 element in streptomyces lividans.	slp1 is a 17.2-kbp genetic element indigenous to the streptomyces coelicolor chromosome. during conjugation, slp1 can undergo excision and subsequent site-specific integration into the chromosomes of recipient cells. we report here the localization, nucleotide sequences, and initial characterization of the genes mediating these recombination events. a region of slp1 adjacent to the previously identified site of integration, attp, was found to be sufficient to promote site-specific integration of ...	1993	8387993
identification of algf in the alginate biosynthetic gene cluster of pseudomonas aeruginosa which is required for alginate acetylation.	mucoid strains of pseudomonas aeruginosa produce a high-molecular-weight exopolysaccharide called alginate that is modified by the addition of o-acetyl groups. to better understand the acetylation process, a gene involved in alginate acetylation called algf was identified in this study. we hypothesized that a gene involved in alginate acetylation would be located within the alginate biosynthetic gene cluster at 34 min on the p. aeruginosa chromosome. to isolate algf mutants, a procedure for loca ...	1993	8394313
expression of the escherichia coli ftsz gene: trials and tribulations of gene fusion studies.	the ftsz gene of escherichia coli, which codes for an essential cell division protein, is subjected to multiple regulation, as shown in part with studies using an ftsz::lacz operon fusion located on phage lambda jfl100. using this same fusion, we sought to isolate regulatory mutants overexpressing ftsz by selecting mutants able to grow on lactose. one lac+ mutant was obtained which overexpressed the ftsz::lacz fusion 70-fold. the mutation responsible for the overexpression lies in a new gene, co ...	1993	8468005
bacterially expressed fabs of monoclonal antibodies neutralizing tumour necrosis factor alpha in vitro retain full binding and biological activity.	antibody fragments specific for the human tumour necrosis factor alpha (tnf alpha) have been cloned from lambda combinatorial expression libraries using total rna obtained from three different hybridoma cell lines of therapeutic interest. the previously described bacteriophage lambda vectors, lambda hc2 and lambda lc1, were modified to create unique antibody cloning sites in the combinatorial construct and a novel tag peptide was inserted at the c-terminal end of the expressed fd chain. sequence ...	1993	8232337
expression and biochemical properties of a protein serine/threonine phosphatase encoded by bacteriophage lambda.	the predicted amino acid sequence encoded by the open reading frame 221 (orf221) of bacteriophage lambda exhibited a high degree of similarity to the catalytic subunits of a variety of protein serine/threonine phosphatases belonging to pp1, pp2a, and pp2b groups. cloning and expression of the orf221 gene in escherichia coli provided direct evidence that the gene codes for a protein serine/threonine phosphatase. the single-subunit recombinant enzyme was purified in soluble form and shown to posse ...	1993	8248155
from the double-helix to novel approaches to the sequencing of large genomes.	elucidation of the structure of dna by watson and crick [nature 171 (1953) 737-738] has led to many crucial molecular experiments, including studies on dna replication, transcription, physical mapping, and most recently to serious attempts directed toward the sequencing of large genomes [watson, science 248 (1990) 44-49]. i am totally convinced of the great importance of the human genome project, and toward achieving this goal i strongly favor 'top-down' approaches consisting of the physical map ...	1993	8276270
the mode of replication is a major factor in segregational plasmid instability in lactococcus lactis.	the effects of the rolling-circle and theta modes of replication on the maintenance of recombinant plasmids in lactococcus lactis were studied. heterologous escherichia coli or bacteriophage lambda dna fragments of various sizes were inserted into vectors based on either the rolling-circle-type plasmid pwv01 or the theta-type plasmid pambeta1. all pambeta1 derivatives were stably maintained. pwv01 derivatives, however, showed size-dependent segregational instability, in particular when large dna ...	1993	16348863
an analogue of the dnaj molecular chaperone in escherichia coli.	escherichia coli dnaj functions as a typical molecular chaperone in coordination with other heat shock proteins such as dnak and grpe in a variety of cellular processes. in this study, it was found that e. coli possesses an analogue of dnaj, as judged from not only its primary structure but also its possible function. this protein, named cbpa (for curved dna-binding protein), was first identified as a dna-binding protein that preferentially recognizes a curved dna sequence. cloning and nucleotid ...	1994	8302830
the gene fimu affects expression of salmonella typhimurium type 1 fimbriae and is related to the escherichia coli trna gene argu.	the gene fimu, located on a recombinant plasmid carrying the salmonella typhimurium type 1 fimbrial gene cluster is closely related to the escherichia coli trna gene argu. the fimu gene complements an e. coli argu mutant that is a p2 lysogen, thereby allowing the phage p4 to grow in this strain but preventing the growth of phage lambda. in addition, fimu was shown to be involved in fimbrial expression since transformants of the e. coli argu mutant could produce fimbriae only in the presence of f ...	1994	7914347
a novel escherichia coli expression-export vector containing alkaline phosphatase as an insertional inactivation screening system.	an escherichia coli expression-export vector was constructed (pcgv1, 6.3 kb) containing the alkaline phosphatase structural gene (phoa) located downstream from the phage lambda pr and pl promoters positioned in tandem and the cits857 gene encoding lambda thermosensitive repressor. the phoa gene is fused to dna encoding a hybrid signal sequence that contains the n-terminal portion of the beta-lactamase (bla) signal sequence and the c-terminal region of the phoa signal sequence. within the dna enc ...	1994	7926833
cloning and expression of the gene for group b streptococcal hyaluronate lyase.	group b streptococci (gbs) are a major cause of serious human perinatal infections. most clinical isolates of gbs secrete hyaluronate lyase, and production of high levels of the enzyme has been associated with strain virulence. degenerate oligonucleotide primers, designed on the basis of the amino acid sequences of tryptic peptides prepared from the purified enzyme, permitted the polymerase chain reaction amplification from gbs chromosomal dna of a 363-base pair internal dna fragment of the gbs ...	1994	7982914
topology of the product binding site in rna polymerase revealed by transcript slippage at the phage lambda pl promoter.	in the presence of transcription substrates atp, ctp, and utp, a stable ternary complex containing tetranucleotide auca is formed on the phage lambda pl promoter (starting sequence c-3a-2c-1a+1u+2c+3a+4g+5). we show that in the absence of gtp or at undersaturating gtp concentrations the auca transcript synthesized at the +1 to +4 segment slips back by 3 nucleotides and is stabilized in the ternary complex in such a way that only its 2 3'-proximal bases remain paired to the -1/+1 positions of the ...	1994	7989343
high-level expression of staphylococcal nuclease a in escherichia coli.	the staphylococcal nuclease a gene has been successfully cloned and overexpressed in e. coli under the transcriptional control of the bacteriophage lambda prpl promoters regulated by the temperature sensitive repressors. the sds-page analysis demonstrates that the nuclease a is produced to the extent of as much as 60% of the total cellular protein. the n-terminal analysis of the nuclease a shows that the amino terminal formyl methionine residue of the enzyme is precisely processed. the recombina ...	1994	7993969
the vibrio cholerae toxin-coregulated-pilus gene tcpi encodes a homolog of methyl-accepting chemotaxis proteins.	virulence gene activation in vibrio cholerae is under the control of the toxr-toxt regulatory cascade. the toxr regulon consists of genes required for toxin-coregulated-pilus (tcp) biogenesis, accessory colonization factor genes, cholera toxin genes, and toxr-activated genes (tag) of unknown function. the tagb gene was isolated by using a tagb::tnphoa fusion junction to probe a v. cholerae )395 bacteriophage lambda library. nucleotide sequence analysis revealed that tagb is identical to tcpi, a ...	1994	8005659
inactivation and repair of bacteriophage lambda by furfural.	furfural is a dietary mutagen and is present in various frequently consumed food products. in earlier work we have shown that it induces single strand breaks in duplex dna which occur preferentially in at base pairs. experiments on the conservation of ecori cleavage site in mutant plasmids further established that repair of furfural damaged plasmid dna occurs on propagation in the host cells. in this paper it is shown that the strand scissions induced by furfural in dna account for its biologica ...	1994	8019442
functions involved in bacteriophage p2-induced host cell lysis and identification of a new tail gene.	successful completion of the bacteriophage p2 lytic cycle requires phage-induced lysis of its escherichia coli host, a process that is poorly understood. genetic analysis of lysis-deficient mutants defined a single locus, gene k, which lies within the largest late transcription unit of p2 and maps between head gene l and tail gene r. we determined and analyzed the dna sequence of a ca. 2.1-kb ecorv fragment that spans the entire region from l to r, thus completing the sequence of this operon. th ...	1994	8051010
[bacteriophage lambda:lux: design and expression of bioluminescence in e. coli cells].	the bacteriophages lambda:lux and lambda:luxab have been constructed by ligation of phage arms generated by bamhi or salgi restriction endonucleases digestion of embl4 to bamhi digested plasmid pf1 lux+ or to salgi digested plasmid pf2 lambda:luxa+b+. cells of escherichia coli prototrophic strain cs were infected with lambda:lux or lambda:luxab and intensity of bioluminiscence of the samples registered at different time intervals determined. the signal of bioluminiscence was first detected 15 mi ...	1994	8065385
synthesis and assembly of the f0 proton channel from f0 genes cloned into bacteriophage lambda and integrated into the escherichia coli chromosome.	the promoter region and the first four genes of the escherichia coli proton-translocating atpase (unc) operon, uncibef, were cloned into bacteriophage lambda, enabling this region to be recombined into an unc-deleted e. coli chromosome at the lambda att site. the resultant e. coli strain, carrying single-copy f0 genes, was tested for synthesis and assembly of functional f0 proton channels. membranes isolated from this strain contained all three f0 subunits and were capable of binding purified f1 ...	1994	8125942
generation of bacteriophage lambda lysogens by electroporation.		1994	8179876
generating bacteriophage lambda sublibraries enriched for rare clones.		1994	8185914
control of transcription processivity in phage lambda: nus factors strengthen the termination-resistant state of rna polymerase induced by n antiterminator.	during transcription of phage lambda early operons, the n gene product alters host rna polymerase (rnap) so that transcription proceeds through multiple stop signals. here, we reproduce the essence of n activity with purified components in synthetic transcription units that contain lambda pl promoter and the n-recognition site, nutl, followed by a variety of intrinsic terminators. we show that three host factors (nusa, nuse, and nusg) are essential for n to allow appreciable transcription throug ...	1994	7521531
differential replication of plasmids during stringent and relaxed response of escherichia coli.	stringent control of dna replication has been demonstrated for a few replicons like oric, pbr322, and plasmids derived from coliphage lambda. in this study we investigated the replication of other plasmids harboring a well defined origin (orip15a, oripsc101, and orirk2 = oriv) in amino-acid-starved stringent and relaxed strains of escherichia coli. we found differential replication of plasmids during stringent and relaxed response. inhibition of dna synthesis or amplification of plasmid dna in a ...	1994	7527562
cloning of the na(+)-translocating nadh-quinone reductase gene from the marine bacterium vibrio alginolyticus and the expression of the beta-subunit in escherichia coli.	the na(+)-translocating nadh-quinone reductase purified from the marine bacterium vibrio alginolyticus is composed of three subunits, alpha, beta and gamma. from the n-terminal amino acid sequences of each subunit and its polypeptide fragment obtained by partial digestion with v8 protease, oligonucleotides corresponding to forward and reverse primers for each gene (nqr a, b and c) encoding the alpha, beta and gamma subunit, respectively, were synthesized. using these primers, a part of each gene ...	1994	7805866
very short patch repair of t:g mismatches in vivo: importance of context and accessory proteins.	in escherichia coli, t:g mismatches in specific contexts are corrected by a very short patch (vsp) repair system. previous studies have shown that the product of gene vsr mediates correction of t:g to c:g in the 5'ctagg/3'ggtcc context and in some related contexts. amber mutations that arose in cag sequences in gene ci of bacteriophage lambda were used to determine the effect of flanking bases on the repair of t:g mispairs arising during phage recombination. the experimental findings were combin ...	1995	7836300
repair of phage lambda dna damaged by near ultraviolet light plus 8-methoxypsoralen.	treatment of phage lambda with 8-methoxypsoralen plus near ultraviolet light (puva) and its subsequent infection and growth on various mutant and non-mutant hosts were investigated. a number of escherichia coli dna repair-deficient mutants, particularly those deficient in genes producing proteins known to participate in interstrand crosslink repair, were used as hosts to assess the roles of these gene products in the activation of phage affected by puva. results show that puva, uvra, uvrd, reca, ...	1995	7897361
a role for a small stable rna in modulating the activity of dna-binding proteins.	the 10sa rna, encoded by the e. coli ssra gene, appears to modulate action of some dna-binding proteins. when ssra is inactivated, lacz expression from the lac operon, as well as galk from a gal operon fused to a phage lambda promoter, is reduced from that observed in bacteria wild-type for ssra. these differences are not observed if the relevant repressor is inactive, suggesting that in the absence of 10sa rna binding of laci and lambda ci repressors is enhanced. gel mobility shifts show that 1 ...	1995	7585940
analysis of the dnak molecular chaperone system of francisella tularensis.	we have cloned the francisella tularensis (ft) grpe-dnak-dnaj heat-shock genes which are organized in that order. these genes allow heterologous genetic complementation of each respective mutant strain of escherichia coli (ec) for bacteriophage lambda growth. the nucleotide sequences of the ft grpe-dnak-dnaj genes and the deduced amino-acid sequences share significant homologies with their respective ec counterparts. the ft dnak and dnaj proteins cross-react with polyclonal antibodies raised aga ...	1995	7590305
the recombination hot spot chi activates recbcd recombination by converting escherichia coli to a recd mutant phenocopy.	the products of the recb and recc genes are necessary for conjugal recombination and for repair of chromosomal double-chain breaks in escherichia coli. the recd gene product combines with the recb and recc proteins to comprise recbcd enzyme but is required for neither recombination nor repair. on the contrary, recbcd enzyme is an exonuclease that inhibits recombination by destroying linear dna. the recd ejection model proposes that recbcd enzyme enters a dna duplex at a double-chain end and trav ...	1995	7603978
rate of translocation of bacteriophage t7 dna across the membranes of escherichia coli.	translocation of bacteriophage t7 dna from the capsid into the cell has been assayed by measuring the time after infection that each gatc site on the phage genome is methylated by cells containing high levels of dna adenine methylase. methylation at gatc sites on t7 dna renders both the infecting genome and any newly synthesized molecules sensitive to the restriction enzyme dpni. in a normal infection at 30 degrees c, translocation of the t7 genome into the cell takes between 9 and 12 min. in co ...	1995	7608081
temperature induction of bacteriophage lambda mutants in escherichia coli.	the paper presents temperature induction in escherichia coli cells with phage lambda on the target-protein production and cell growth. replicated lambda-dna particles in the q- and s- mutants remain naked for a longer time by preventing dna packaging and cell lysis, and therefore the expression of the foreign genes is high. however, the parasitic infection of phage-lambda causes on significant losses of host cell viability in the induction phase. the temperature effects on cell growth and target ...	1995	7612243
gene cloning, sequence analysis, purification, and secretion by escherichia coli of an extracellular lipase from serratia marcescens.	the gene encoding extracellular lipase of serratia marcescens has been identified from a phage lambda genomic library. formation of orange-red fluorescent plaques on rhodamine b-triolein plates was used to identify phages carrying the lipase gene. a 2.8-kb sali fragment was subcloned into a plasmid, and lipase was expressed in escherichia coli. extracellular lipase was detected in the presence of the secretion plasmid pgsd6 carrying the genes prtd, -e, and -f, which guide the secretion of protea ...	1995	7618881
expression, purification and characterisation of the product from the bacillus subtilis hemd gene, uroporphyrinogen iii synthase.	uroporphyrinogen iii synthase, the product of the hemd gene, is the enzyme responsible for the cyclisation of the linear tetrapyrrole, hydroxymethylbilane. the hemd gene isolated from bacillus subtilis was manipulated by pcr to enable direct cloning behind a synthetic ribosome-binding site downstream of tandem bacteriophage lambda pr and pl promoters in a pce30-derived vector. following thermal induction of transcription, the resulting plasmid (pps21) directed the synthesis of uroporphyrinogen i ...	1995	7628476
the rhizobium meliloti groelc locus is required for regulation of early nod genes by the transcription activator nodd.	the molecular chaperones related to groel (hsp60, cpn60) interact with partially folded proteins and appear to assist them to attain active and correctly folded conformation. they are required for cell viability but are probably more important for some processes than for others. through a random genetic search to find loci that are required for expression of the rhizobium meliloti nod (nodulation) genes, we isolated a mutant (b4) defective in luteolin-dependent activation of nod gene expression, ...	1995	7729688
the requirement for molecular chaperones in lambda dna replication is reduced by the mutation pi in lambda p gene, which weakens the interaction between lambda p protein and dnab helicase.	during the initiation of lambda dna replication, the host dnab helicase is complexed with phage lambda p protein in order to be properly positioned near the ori lambda-lambda o initiation complex. however, the lambda p-dnab interaction inhibits the activities of dnab. thus, the concerted action of bacterial heat shock proteins, dnak, dnaj, and grpe, is required to activate the helicase. wild-type phage lambda cannot grow on the e. coli dnab, dnak, dnaj, and grpe mutants. however, lambda phage wi ...	1995	7730358
cloning and expression of cdna encoding a protein that binds a palindromic promoter element essential for induction of fungal cutinase by plant cutin.	previous studies showed that a palindromic sequence located at -159 base pairs is essential for induction of cutinase gene in fusarium solani f. sp. pisi (nectria haematococca mating type vi) by the hydroxy fatty acids from plant cutin and that a 50-kda nuclear protein binds to a promoter that contains this element. screening of a phage lambda gt11 expression library with the concatenated palindromic sequence as the probe identified a cdna encoding a palindrome-binding protein (pbp). nucleotide ...	1995	7744822
production of a biologically active recombinant teleostean growth hormone in e. coli cells.	we have isolated and characterized several recombinant lambda phage clones carrying growth hormone (gh) cdna of striped bass (morone saxatilis). nucleotide sequence and the predicted amino acid sequence of sbgh was determined from a recombinant clone carrying the longest cdna insert. the sbgh cdna encodes a pre-hormone of 204 amino acid residues. comparison of the predicted amino acid sequence of sbgh with those of other vertebrates revealed different degrees of sequence identity: approximately ...	1995	7758842
phage t4 dna [n6-adenine]methyltransferase. overexpression, purification, and characterization.	the bacteriophage t4 dam gene, encoding the dam dna [n6-adenine]methyltransferase (mtase), has been subcloned into the plasmid expression vector, pjw2. in this construct, designated pint4dam, transcription is from the regulatable phage lambda pr and pl promoters, arranged in tandem. a two-step purification scheme using deae-cellulose and phosphocellulose columns in series, followed by hydroxyapatite chromatography, was developed to purify the enzyme to near homogeneity. the yield of purified pro ...	1995	7782299
glms of thermus thermophilus hb8: an essential gene for cell-wall synthesis identified immediately upstream of the s-layer gene.	a 30 kbp chromosomal region containing the s-layer gene (slpa) from thermus thermophilus hb8 was cloned from a lambda phage gene library. dna sequence analysis of the region upstream to the slpa gene revealed the presence of an open reading frame (orf) which coded for a 604-amino-acid protein highly homologous to the glucosamine-6-p synthases (ec 2.6.1.16) of both prokaryotic and eukaryotic origin. the identification of this orf as the glucosamine-6-p synthase gene from t. thermophilus (glmsth) ...	1995	7476196
biosynthetic incorporation of 7-azatryptophan into the phage lambda lysozyme: estimation of tryptophan accessibility, effect on enzymatic activity and protein stability.	the phage lambda lysozyme (lambda l) contains four tryptophans. these have been efficiently replaced by 7-azatryptophan (7aw) through biosynthetic incorporation into the overexpressed protein. comparative analysis of the effect of temperature or ph on the fluorescence of the wild-type lambda l and 7aws-containing protein (a lambda l) shows that the stability of the protein is only mildly reduced by 7aw incorporation above ph 5 but that it is strongly decreased below ph 4 on protonation of inacce ...	1995	8532666
functional analysis of isolated cpn10 domains and conserved amino acid residues in spinach chloroplast co-chaperonin by site-directed mutagenesis.	the possibilities of independent function of the two chaperonin 10 (cpn10) domains of the cpn10 homologue from spinach chloroplasts and the role of five conserved amino acid residues in the n-terminal cpn10 unit were investigated. recombinant single domain proteins and complete chloroplast cpn10 proteins carrying amino acid exchanges of conserved residues in their n-terminal cpn10 domain were expressed in escherichia coli and partially purified. the function of the recombinant proteins was teste ...	1995	8555447
transformation of naturally competent streptococcus mutans with replicative and non-replicative tn916-containing plasmids: implications for a mechanism of transposition.	based on the observations reported here and what is known concerning transformation of naturally competent strains of s. mutans and other streptococcal species such as s. gordonii, we propose the model shown in figure 2. the tn916-intermediate transforms s. mutans as originally proposed for b. subtilis by scott and coworkers [8]. it is not clear in either system (b. subtilis or s. mutans) whether the tn916 intermediate enters the cell as ds-dna or ss-dna. because it is likely that transformation ...	1995	8586174
transcription antitermination: the lambda paradigm updated.	coliphage lambda employs systems of transcription termination and antitermination to regulate gene expression. early gene expression is regulated by the phage-encoded n protein working with a series of escherichia coli proteins, nus, at rna sites, nut, to modify rna polymerase to a termination-resistant form. expression of lambda late genes is regulated by the phage-encoded q antitermination protein. q, which appears to use only one host factor, acts at a dna site, qut, to modify rna polymerase ...	1995	8709839
function of e. coli rna polymerase sigma factor sigma 70 in promoter-proximal pausing.	the sigma factor sigma 70 of e. coli rna polymerase acts not only in initiation, but also at an early stage of elongation to induce a transcription pause, and simultaneously to allow the phage lambda gene q transcription antiterminator to act. we identify the signal in dna that induces early pausing to be a version of the sigma 70 -10 promoter consensus, and we show that sigma 70 is both necessary for pausing and present in the paused transcription complex. regions 2 and 3 of sigma 70 suffice to ...	1996	8756730
[organization of the gene for the beta-subunit of human photoreceptor cyclic gmp phosphodiesterase].	two recombinant bacteriophage lambda clones encoding a 27.8-kb fragment of the human phosphodiesterase beta-subunit gene were isolated from a human genomic library. the nucleotide sequences of 19 exons (from the 4th to 22nd), 18 introns, and the 3'-flanking region were determined. the analysis of the nucleotide sequence of the phosphodiesterase beta-subunit gene revealed four alu repeats and four minisatellite regions.	1996	8768262
device for the simultaneous screening of bacteriophage lambda clones.		1996	8770396
conjugative transposon tn916: evidence for excision with formation of 5'-protruding termini.	conjugative transposons are genetic elements able to promote their own intracellular transposition and intercellular conjugal transfer. they move by an excision-integration system related to that of lambdoid phages, in which the first step is the excision of the transposon from the donor replicon to form a covalently closed circular intermediate which contains a heteroduplex joint. in this work, sequencing both strands of the circular intermediate heteroduplex joint, it was found that, as during ...	1996	8824634
modulation of lambda integrase synthesis by rare arginine trna.	lambda's int gene contains an anomalously high frequency of the rare arginine codons aga and agg when compared to genes of escherichia coli or to the rest of phage lambda. these are the least frequent codons in genes of e. coli and are recognized by the rarest trnas. the presence of these codons reduces the translation rate and, depending on the context, this can strongly modulate translational efficiency by a variety of mechanisms. in this study, we show that expression of the natural int gene ...	1996	8843435
identification and characterization of the escherichia coli rbn gene encoding the trna processing enzyme rnase bn.	the gene encoding rnase bn was localized to 88 min on the escherichia coli chromosome by a novel suppressor assay and conjugational and transductional analysis. assay of subclones derived from lambda phage 543 of the kohara library, which encompasses this region of the chromosome, for elevated rnase bn activity identified o290, a previously reported open reading frame, as the gene encoding rnase bn. interruption of this gene with a kan(r) cassette and introduction into the chromosome eliminated ...	1996	8955422
[lux-regulon from vibrio fisheri reduces type 1 restriction in escherichia coli k12 cells].	the ecok restriction of nonmodified phage lambda.0 controlled by escherichia coli k12 lon- cells is alleviated 10-20-fold in the presence of a hybrid plasmid containing the entire lux regulon of vibrio fischeri. the la protease participates in degradation of the regulatory luxr-luxi proteins; therefore, transcription of the right lux operon begins significantly earlier than in the lon+ bacteria and is characterized by high content of the n-(3-oxohexanoyl) homoserine lactone autoinducer in medium ...	1996	8964486
role of escherichia coli rna polymerase alpha subunit in modulation of pausing, termination and anti-termination by the transcription elongation factor nusa.	the alpha subunit (alpha) of rna polymerase (rnap) is critical for assembly of polymerase and positive control of transcription initiation in escherichia coli. here, we report that alpha also plays a role in transcription elongation, and this involves a direct interaction between alpha and nusa factor. during in vitro transcription without nusa, alpha interacts with the nascent rna, as revealed by photocrosslinking. when nusa is present, rna crosslinks to nusa rather than to alpha. we show that ...	1996	8598198
long-circulating bacteriophage as antibacterial agents.	the increased prevalence of multidrug-resistant bacterial pathogens motivated us to attempt to enhance the therapeutic efficacy of bacteriophages. the therapeutic application of phages as antibacterial agents was impeded by several factors: (i) the failure to recognize the relatively narrow host range of phages; (ii) the presence of toxins in crude phage lysates; and (iii) a lack of appreciation for the capacity of mammalian host defense systems, particularly the organs of the reticuloendothelia ...	1996	8622911
escherichia coli thymidylate kinase: molecular cloning, nucleotide sequence, and genetic organization of the corresponding tmk locus.	thymidylate kinase (dtmp kinase; ec 2.7.4.9) catalyzes the phosphorylation of dtmp to form dtdp in both de novo and salvage pathways of dttp synthesis. the nucleotide sequence of the tmk gene encoding this essential escherichia coli enzyme is the last one among all the e. coli nucleoside and nucleotide kinase genes which has not yet been reported. by subcloning the 24.0-min region where the tmk gene has been previously mapped from the lambda phage 236 (e9g1) of the kohara e. coli genomic library ...	1996	8631667
identification of functional regions of the nun transcription termination protein of phage hk022 and the n antitermination protein of phage lambda using hybrid nun-n genes.	phages lambda and hk022 express proteins n and nun, respectively, each of which acts with a number of escherichia coli host nus factors at lambda nut rna sites, to influence transcription elongation. the lambda nut sites, nearly identical sequences located downstream of the early promoters, pl and pr, were first identified as cis-acting signals required for the action of n in forming termination-resistant transcription complexes. surprisingly, the nun protein, resembling n and expressed by anoth ...	1996	8632463
the rna chain elongation rate of the lambda late mrna is unaffected by high levels of ppgpp in the absence of amino acid starvation.	in this study, the effects of high levels of guanosine tetraphosphate (ppgpp) on the decay and rna chain elongation kinetics of the bacteriophage lambda late transcript in escherichia coli were examined in the absence of amino acid starvation. the accumulation, mrna decay kinetics, and rna chain elongation rate of the lambda late mrna were determined after heat induction of lambdaci857 lysogens in the presence of high levels of ppgpp induced from a relaalpha fragment-overproducing plasmid. the a ...	1996	8663373
rho-dependent termination of transcription is governed primarily by the upstream rho utilization (rut) sequences of a terminator.	a rho-dependent transcription terminator in escherichia coli dna consists of an upstream part for rho utilization (rut) and the transcription stop point (tsp) region. to test the role of the tsp region variants of the coliphage lambda cro gene terminator, tr1, containing inserts of non-terminator sequences between its rut and tsp regions were tested for termination function. the results showed that termination occurred with high efficiency at multiple sites in each of the new sequences with the ...	1996	8702947