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L Daneo-Moore

Publications and source records attributed to L Daneo-Moore.

At least 19 recordsLinked to original sources
PubMed1992

Modular design of the Enterococcus hirae muramidase-2 and Streptococcus faecalis autolysin.

The mature forms of the extracellular muramidase-2 of Enterococcus hirae and Streptococcus faecalis autolysin have very similar primary structures. Each consists of an active-site-containing N-terminal domain fused to a multiple-repeat C-terminal domain. Polypeptide segments occurring at equivalent places in these two bacterial wall lytic enzymes have homologues in two phage lysozymes and in three functionally unrelated proteins, illustrating the principle that protein molecules frequently are constructed from modules that are linked in a single polypeptide chain.

Amino Acid Sequence↗
PubMed1992

A truncated Tn916-like element in a clinical isolate of Enterococcus faecium.

A 58.7-kb nonconjugative plasmid (pKQ1) previously reported in a clinical isolate of Enterococcus faecium was found to contain both a tetM and an erythromycin resistance (erm) determinant. The plasmid contained a region homologous to the A, F, H, and G HincII fragments of Tn916. However, the 4.8-kb B fragment of Tn916 which contained the tetM determinant was replaced by a 7.3-kb fragment, and the 3.6-kb HincII C fragment of Tn916 was missing. An element homologous to Tn917 was juxtaposed to the truncated Tn916-like element. The Tn917-like element was similar in size to the erm transposon Tn917 as determined by a ClaI restriction digest which spanned approximately 99% of the transposon. When Bacillus subtilis or Streptococcus sanguis were transformed with pKQ1, no zygotically induced transposition of the tetM element was detected. Similarly no transposition of the Tn917-like element was detected.

Bacillus subtilis↗
PubMed1992

Cloning and sequence analysis of the muramidase-2 gene from Enterococcus hirae.

Extracellular muramidase-2 of Enterococcus hirae ATCC 9790 was purified to homogeneity by substrate binding, guanidine-HCl extraction, and reversed-phase chromatography. A monoclonal antibody, 2F8, which specifically recognizes muramidase-2, was used to screen a genomic library of E. hirae ATCC 9790 DNA in bacteriophage lambda gt11. A positive phage clone containing a 4.5-kb DNA insert was isolated and analyzed. The EcoRI-digested 4.5-kb fragment was cut into 2.3-, 1.0-, and 1.5-kb pieces by using restriction enzymes KpnI, Sau3AI, and PstI, and each fragment was subcloned into plasmid pJDC9 or pUC19. The nucleotide sequence of each subclone was determined. The sequence data indicated an open reading frame encoding a polypeptide of 666 amino acid residues, with a calculated molecular mass of 70,678 Da. The first 24 N-terminal amino acids of purified extracellular muramidase-2 were in very good agreement with the deduced amino acid sequence after a 49-amino-acid putative signal sequence. Analysis of the deduced amino acid sequence showed the presence at the C-terminal region of the protein of six highly homologous repeat units separated by nonhomologous intervening sequences that are highly enriched in serine and threonine. The overall sequence showed a high degree of homology with a recently cloned Streptococcus faecalis autolysin.

Amino Acid Sequence↗
PubMed1990

Size of the Streptococcus mutans GS-5 chromosome as determined by pulsed-field gel electrophoresis.

Rare cutting restriction endonucleases were used to cut the Streptococcus mutans chromosome into large fragments. Restriction enzymes utilizing recognition sites containing 6-, 7-, or 8-base-pair sequences with only G and C nucleotides produced few fragments, most of which were greater than 100 kilobase pairs in size. Addition of the fragments from digests of SmaI, NotI, ApaI, RsrII, and EagI yielded a molecular size for the S. mutans GS-5 genome of 2,819 +/- 60 kilobase pairs.

Chromosomes, Bacterial↗
PubMed1990

Properties of cell wall-associated DD-carboxypeptidase of Enterococcus hirae (Streptococcus faecium) ATCC 9790 extracted with alkali.

DD-Carboxypeptidase (DD-CPase) activity of Enterococcus hirae (Streptococcus faecium) ATCC 9790 was extracted from intact bacteria and from the insoluble residue (crude cell wall fraction) of mechanically disrupted bacteria by a brief treatment at pH 10.0 (10 mM glycine-NaOH) at 0 degrees C or by extraction with any of several detergents. Extractions with high salt concentrations failed to remove DD-CPase activity from the crude wall fraction. In contrast to N-acetylmuramoylhydrolase (both muramidase 2 and muramidase 1) activities, DD-CPase activity failed to bind to insoluble cell walls or peptidoglycan matrices. Thus, whereas muramidase 1 and muramidase 2 activities can be considered to be cell wall proteins, the bulk of the data are consistent with the interpretation that the DD-CPase of this species is a membrane protein that is sometimes found in the cell wall fraction, presumably because of hydrophobic interactions with other proteins and cell wall polymers. The binding of [14C]penicillin to penicillin-binding protein 6 (43 kilodaltons) was proportional to DD-CPase activity. Kinetic parameters were also consistent with the presence of only one DD-CPase (penicillin-binding protein 6) in E. hirae.

Cell Membrane↗
PubMed1989

Transposon-916-like elements in clinical isolates of Enterococcus faecium.

Tetracycline (Tc) resistance was found in nine out of ten clinical isolates of Enterococcus faecium. Conjugative transposons, designated Tn5031, Tn5032 and Tn5033, were present in the chromosome of three isolates. The transposons were similar both structurally and functionally to Tn916 containing the tetM determinant. A large non-conjugative plasmid found in a fourth isolate contained an element homologous to Tn916. The four isolates containing the element showing homology to Tn916 exhibited a substantially higher level of Tc resistance than the remaining five Tc-resistant isolates. Tc-resistance genes which have not been identified are apparently responsible for the low-level Tc resistance in five clinical isolates.

Conjugation, Genetic↗
PubMed1989

Effect of cell cycle stages on the central density of Enterococcus faecium ATCC 9790.

Cultures of Enterococcus faecium growing at various rates were examined for timing of cell division cycle events by using the method of residual divisions and a morphological analysis. Both methods gave essentially the same timing for the onset of D1 (completion of chromosome replication) and of D2 (completion of septation). Frequencies of cells exhibiting a phase-reversed center in bovine serum albumin at various growth rates were determined. The data fit a model in which rapidly growing cells increase in refractive index (which is assumed to represent central density) at completion of the chromosome replication cycle involved in the ongoing division, whereas slowly growing cultures increase in central density at the time of completion of septation. There was no correlation between the timing of increase in central density and the timing of initiation of new sites of surface growth.

Cell Cycle↗
PubMed1989

The second peptidoglycan hydrolase of Streptococcus faecium ATCC 9790 covalently binds penicillin.

A second peptidoglycan hydrolase (muramidase-2) of Streptococcus faecium ATCC 9790 (Enterococcus hirae) has been purified to apparent homogeneity. The enzyme has been shown to be a beta-1,4-N-acetylmuramoylhydrolase (muramidase; EC 3.2.1.17) and to differ in substrate specificity from a previously isolated muramidase. Purified enzyme appears as two protein staining bands with molecular masses of 125 and 75 kilodaltons (kDa) on polyacrylamide gels after sodium dodecyl sulfate electrophoresis. Elution and renaturation of protein bands from sodium dodecyl sulfate-polyacrylamide gels showed that both proteins have muramidase-2 activity. Both proteins have been shown to bind radioactive benzylpenicillin and have the same electrophoretic mobilities as penicillin-binding proteins 1 and 5 present in membrane preparations of this organism, respectively. Incubation of a [14C]penicillin G-labeled 125-kDa form of the enzyme with crude alkaline extracts from S. faecium (which did not contain added proteinase inhibitors) showed the endogenous conversion of the radiolabeled 125-kDa form to the radiolabeled 75-kDa form of the enzyme.

Chromatography, Affinity↗
PubMed1988

Transformation of Streptococcus sanguis to intrinsic penicillin resistance.

A series of step-level penicillin-resistant derivatives of Streptococcus sanguis V288 (Challis) were obtained through successive genetic transformations. The DNA donor used was a laboratory-derived, penicillin-resistant multistep mutant of the recipient strain. Detection of the penicillin-binding proteins (PBPs) of wild-type and transformants revealed five major PBPs. While it was found that S. sanguis can acquire intrinsic resistance in a stepwise manner and the mechanism was similar to those of some other organisms (changes in penicillin-binding protein affinity and/or in extent of penicillin binding), multiple-PBP changes accompanied a single step-level of resistance. All of the PBPs showed varying degrees of decreased affinity for [3H]benzylpenicillin with increasing penicillin resistance. Of these, the consistent, dramatic and progressive decrease of PBP 4 binding was most notable. After an initial decrease at the first step-level of resistance, PBP 5 was restored to wild-type levels, indicating a possible important role in survival. Genetic linkage of the first two step-levels of resistance was demonstrated by examination of transformation frequencies and by hit-kinetics experiments. A convenient method is described for the quantitative comparison of fluorographs containing PBPs with a wide range of affinities for penicillin.

Bacterial Proteins↗
PubMed1988

Tn916 insertional inactivation of multiple genes on the chromosome of Streptococcus mutans GS-5.

Streptococcus mutans GS-5 was transformed with the Escherichia coli plasmid pAM150 containing the cloned streptococcal transposon Tn916. Southern blot analyses with the tetracycline-resistant determinant of Tn916 showed that Tn916 was inserted into the chromosome of S. mutans at a variety of different sites. Tn916 insertions resulted in the inactivation of genes that code for various steps in the biosynthesis of several different amino acids. Two auxotrophs which contained a single copy of Tn916 were shown to revert to prototrophy at frequencies of about 10(-8). All of the revertant prototrophs were susceptible to tetracycline, indicating regeneration of the functional gene by excision of Tn916.

DNA Transposable Elements↗
PubMed1987

Penicillin tolerance in Streptococcus faecium ATCC 9790.

Tolerant strains of Streptococcus faecium had higher levels of muramidase 2 and lower levels of trypsinactivable muramidase 1 than did susceptible strains. Susceptible strains lysed faster than did tolerant strains in buffer and at some antibiotic concentrations. The addition of Triton X-100 produced equal lysis rates for susceptible and tolerant cultures.

Bacteriolysis↗
PubMed1986

Inhibition of beta-lactam antibiotics at two different times in the cell cycle of Streptococcus faecium ATCC 9790.

Treatment of Streptococcus faecium ATCC 9790 with sublytic concentrations of beta-lactam antibiotics revealed two different division blocks in the cell division cycle. One block, induced by N-formimidoyl thienamycin and methicillin, occurred before the completion of chromosome replication, whereas the other, induced by cefoxitin and cephalothin, took place later in the cycle. In addition, these antibiotics gave rise to distinct morphological forms; the antibiotics acting at the earlier block point produced mainly "dumbbells," whereas those affecting the later time formed "lemons." When used in combination N-formimidoyl thienamycin and cefoxitin exerted synergistic killing on this strain. These data suggest that beta-lactam antibiotics have at least two sites of action in S. faecium.

Anti-Bacterial Agents↗
PubMed1986

Relationship of shape to initiation of new sites of envelope growth in Streptococcus faecium cells treated with beta-lactam antibiotics.

Exponential-phase cells of Streptococcus faecium were treated with concentrations of ampicillin and cephalothin which, over 60 min, had little effect on increase in culture mass but resulted in about a 65% inhibition of increase in cell numbers. The resulting drug-treated cells underwent about a doubling in cell mass and volume above that of the untreated cells. The newly divided cells produced in the presence of drugs were shown to be due to the division of central or primary sites of envelope growth present at the time of treatment. Sites that were newly initiated (secondary sites) at the time of treatment or sites initiated after treatment did not divide but enlarged in length and girth to give abnormally large cells. Although the increase in average total volume was the same after each interval of treatment with ampicillin and cephalothin, the primary growth sites of the cephalothin-treated cells grew somewhat more slowly, and their secondary sites grew somewhat more quickly, than did those of the ampicillin-treated cells. Cephalothin-treated cells initiated secondary sites at a rate similar to that of the untreated cells, whereas the ampicillin-treated cells exhibited reduced rates of secondary site initiation. Two models are presented that account for these results.

Ampicillin↗
PubMed1986

Autoradiographic studies of the synthesis of RNA and protein as a function of cell volume in Streptococcus faecium.

Mid-exponential-phase cultures were either labeled continuously with tritiated leucine and uracil or pulse-labeled with tritiated leucine. The amount of leucine and uracil incorporated into protein or RNA per cell was determined by grain counts of autoradiographs of cells seen in electron micrographs; the volume of each cell was determined by three-dimensional reconstruction. The average number of autoradiographic grains around cells continuously labeled with uracil and leucine increased linearly with cell volume. In contrast, while the average grain count around cells pulse-labeled with leucine increased in a near-linear fashion over most of the volume classes, less than the expected number of grains were seen around cells in large- and small-size classes. The distribution of grains around cells from both the continuously and pulse-labeled populations could be fit at the 5% confidence level with a Poisson distribution modified to take into consideration the volume distribution of each population of cells analyzed. These findings suggested that large changes in the density of RNA and protein do not occur in most cells as they increase in size; however, there may be decreases in the rate of protein synthesis in some large and small cells. The decrease in the rate of protein synthesis appears consistent with the hypothesis that new sites of envelope growth must be introduced into cells that are close to the division event to restore rapid growth.

Autoradiography↗
PubMed1986

Autoradiographic studies of chromosome replication during the cell cycle of Streptococcus faecium.

Analysis of the distribution of autoradiographic grains around cells of Streptococcus faecium which had been either continuously or pulse-labeled with tritiated thymidine (mass doubling time, 90 min) showed a non-Poisson distribution even when the distribution of cell sizes in the populations studied was taken into account. These non-Poisson distributions of grains were assumed to reflect the discontinuous nature of chromosome replication. To study this discontinuous process further, we fitted an equation to the grain distribution observed for the pulse-labeled cells that assumed that in any population of cells there were subpopulations in which there were zero, one, or two replicating chromosomes. This analysis predicted an average time for chromosome replication and for the period between completion of rounds of chromosome replication and division of 55 and 43 min, respectively, which were in excellent agreement with estimates made by other techniques. The present investigation extended past studies in indicating that the initiation and completion of rounds of chromosome replication are poorly phased with increases in cell volume and that the amount of chromosome replication may be different in different cell halves.

Autoradiography↗
PubMed1985

Recombination-deficient Streptococcus sanguis.

A UV-sensitive derivative was obtained from Streptococcus sanguis Challis. The organism could be transformed with a number of small streptococcal plasmids at frequencies equal to, or 1 logarithm below, the transformation frequencies for the parent organism. However, transformation with chromosomal DNA was greatly impaired in the UV-sensitive derivative.

Methyl Methanesulfonate↗
PubMed1979

Effect of growth rate on lipid and lipoteichoic acid composition in Streptococcus faecium.

The lipid composition of Streptococcus faecium (S. faecalis ATCC 9790) was analyzed at various growth rates. Diphosphatidylglycerol and the non-ionic lipid fraction containing diacylglycerols and neutral glycolipids appeared to accumulate relative to cellular mass as the culture mass doubling time increased from 30 to 80 min. Within the same range of doubling times the non-ionic lipid fraction appeared to become substantially enriched with diacylglycerols. All lipid species and cellular lipoteichoic acid accumulated relative to the cellular mass at doubling times exceeding 80 min, although diacylglycerol accumulation exceeded that of all other compounds studied.

Enterococcus faecalis↗