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A J Zuccarelli

Publications and source records attributed to A J Zuccarelli.

12 recordsLinked to original sources
PubMed1997

The Aer protein and the serine chemoreceptor Tsr independently sense intracellular energy levels and transduce oxygen, redox, and energy signals for Escherichia coli behavior.

We identified a protein, Aer, as a signal transducer that senses intracellular energy levels rather than the external environment and that transduces signals for aerotaxis (taxis to oxygen) and other energy-dependent behavioral responses in Escherichia coli. Domains in Aer are similar to the signaling domain in chemotaxis receptors and the putative oxygen-sensing domain of some transcriptional activators. A putative FAD-binding site in the N-terminal domain of Aer shares a consensus sequence with the NifL, Bat, and Wc-1 signal-transducing proteins that regulate gene expression in response to redox changes, oxygen, and blue light, respectively. A double mutant deficient in aer and tsr, which codes for the serine chemoreceptor, was negative for aerotaxis, redox taxis, and glycerol taxis, each of which requires the proton motive force and/or electron transport system for signaling. We propose that Aer and Tsr sense the proton motive force or cellular redox state and thereby integrate diverse signals that guide E. coli to environments where maximal energy is available for growth.

Amino Acid Sequence↗
PubMed1995

A general method for detecting and sizing large plasmids.

We have devised a method for detecting and estimating the sizes of large bacterial plasmids in the presence of genomic DNA by pulsed-field gel electrophoresis (PFGE). Bacteria harboring plasmids were embedded in agarose and lysed using a rapid protocol. Plugs were incubated with S1 nuclease and subjected to PFGE in agarose gels. S1 nuclease converted supercoiled plasmids into full-length linear molecules. Large plasmids migrated as discrete bands that were readily observed after ethidium staining. Their sizes were reliably estimated by comparison with linear DNA markers. Without S1 digestion, supercoiled plasmids migrated at rates that were not a simple function of their molecular weights, making size determinations problematic. S1-PFGE detected megaplasmids up to 609 kilobases (kb) in six genera of bacteria (Agrobacterium, Escherichia, Klebsiella, Pseudomonas, Salmonella, and Staphylococcus). The procedure gave size values consistent with previous estimates for characterized megaplasmids. Eight new plasmids between 102 and 316 kb were discovered in Klebsiella and Staphylococcus. S1-PFGE avoids the difficulties of plasmid isolation, eliminates the preparation of probes, and does not require knowledge of restriction enzyme cleavage sites. It detects multiple large plasmids up to the limits of PFGE and can be used to screen for megaplasmids in many strains simultaneously.

Bacteria↗
PubMed1991

Salmonella phage PSP3, another member of the P2-like phage group.

Freshly isolated DNA of phage PSP3, whose morphology closely resembles that of phage P2, contained both circular and linear molecules about 31 kb in length. Linear PSP3 DNA molecules possess single-stranded cohesive termini (cos). Sequencing of the fragment anticipated to contain cos revealed a 19-base sequence identical to cos of phage 186. Of the 107 bp to the right of cos, 94 were identical in 186 DNA (88% similarity), and of the 370 bp to the left, 229 were identical (62% similarity). Cos flanking sequences in both P2 and P4 were also highly conserved in PSP3. A number of restriction sites were at similar locations on the two phage DNAs. The parasitic phage P4 propagated on PSP3 lysogens. PSP3 integrates into the Escherichia coli chromosome at 27 min.

Base Sequence↗
PubMed1990

Diversity and stability of restriction enzyme profiles of plasmid DNA from methicillin-resistant Staphylococcus aureus.

Nosocomial infections caused by methicillin-resistant Staphylococcus aureus (MRSA) are a significant epidemiological problem. Detecting the sources of epidemic strains and preventing their access to patients, however, depend upon the availability of techniques to reliably distinguish among MRSA strains. We evaluated restriction enzyme analysis of plasmid DNA for use as an epidemiological marker of MRSA strains. The diversity of plasmid types was assessed by examining 120 clinical and environmental MRSA isolates from five southern California hospitals and from the American Type Culture Collection. Thirty-seven distinctive EcoRI digestion patterns were observed. We characterized each strain by the number of plasmids it contained and the sizes of the fragments that were generated by EcoRI. Very few of the isolates (4.2%) lacked plasmids, and some (6.7%) contained DNA that was not digested by EcoRI. Several isolates (12.5%) contained two or more plasmids. We were able to assess the stability of MRSA plasmid types by tracking epidemic strains over a 2-year period. We also examined successive isolates from 10 individual patients during their hospitalization. In all but one case, the patient's plasmid profiles remained unchanged. We conclude that the diversity and stability of MRSA plasmid types make them excellent epidemiological markers. In support of this conclusion, we found that our data provided significant epidemiological insights. Two epidemic strains, accounting for more than half of the infections, were identified in the five hospitals. The remaining cases were sporadic, caused by MRSA strains that appeared very infrequently and that may have originated from sources outside the hospitals.

Cross Infection↗
PubMed1989

Rapid isolation of high-molecular-weight DNA from agarose gels.

We have developed a simple, reliable, and rapid method for recovering DNA from agarose gels. While many methods for DNA extraction have already been described, few provide quantitative recovery of large DNA molecules. These procedures generally require costly apparatus, extended elution times, or considerable handling of the sample after elution. Our method employs a novel electroelution chamber constructed from acrylic plastic. Gel slices containing DNA are placed in the chamber between platinum electrodes. Voltage is applied and a continuous flow of buffer sweeps the eluted DNA from the chamber into an external receptacle. Elution is complete in 7 min. Concentrated DNA is obtained by butanol extraction and alcohol precipitation in 1 h. Recoveries, quantitated by counting radiolabeled DNA or by densitometry of analytical gels, were 94 to 100% for fragments of 4 to 50 kb. The eluted DNA was undegraded and could be digested with restriction enzymes, ligated, end-labeled, or used to transform cells as efficiently as noneluted DNA. Complete elution of a 100-kb plasmid, a 194-kb concatemer of bacteriophage lambda, and of 440- and 550- chromosomes of Saccharomyces cerevisiae was also achieved using the same process. This method is suitable for routine use in a wide range of cloning applications, including the electrophoretic isolation of large DNA molecules.

DNA↗
PubMed1975

Recombinant DNA molecules of bacteriophage phi chi174.

Phi chi174 DNA structures containing two different parental genomes were detected genetically and examined by electron microscopy. These structures consisted of two monomeric double-stranded DNA molecules linked in a figure 8 configuration. Such DNA structures were observed to be formed preferentially in host recA+ cells or recA+ cell-free systems. Since the host recA+ allele is required for most phi chi174 recombinant formation, we conclude that the observed figure 8 molecules are intermediates in, or end products of, 1 phi chi174 recombination event. We propose that recombinant figure 8 DNA molecules arise as a result of "single-strand aggression," are stabilized by double-strand "branch migration," and represent a specific example of a common intermediate in genetic recombination.

Cell-Free System↗
PubMed1974

Genetic recombination in bacteriophage phi chi 174.

Genetic recombination in bacteriophage phiX174 usually takes place early in the infection process and involves two parental replicative form (double-stranded) DNA molecules. The host recA protein is required; none of the nine known phiX174 cistron products is essential. The products of a single recombination event are nonreciprocal and asymmetric. Typically, only one of the parental genotypes and one recombinant genotype are recovered from a single cell. An alternative, less efficient recombination mechanism which requires an active phiX174 cistron A protein is observed in the absence of the host recA gene product.

Alleles↗
PubMed1972

Deletion mutants of bacteriophage phiX174.

Mutants of bacteriophage varphiX174 have been isolated that are less dense than wild-type varphiX particles in CsCl. When mutant viral (+) strand DNA and wild-type complementary (-) strand DNA are hybridized, the resulting duplex molecules have single-stranded loops characteristic of wild-type-deletion heteroduplexes. The mutant bacteriophages fail to complement varphiX amber mutants in cistron E. We conclude that the mutant viruses have deleted approximately 7% of the varphiX genome in the region of cistron E.

Carbon Isotopes↗
PubMedDate not supplied

Microbiologic evaluation of a hydrogen peroxide sterilization system.

The reliability of chemical sterilizers (acetone and/or 30-percent hydrogen peroxide at 25 degrees C and at 60 degrees C) was tested against Bacillus subtilis inoculated onto glass slides, commercial biological indicator discs (Bacillus stearothermophilus and B. subtilis), and B. subtilis spore survival. Acetone alone was not sporicidal. Hydrogen-peroxide-sterilized glass slides were sterile after 5 minutes. The indicator discs required 25 minutes at 25 degrees C, and less than 3 minutes at 60 degrees C (P < .0001). The D value of B. subtilis in 27-percent hydrogen peroxide at 25 degrees C is 2 minutes, with z values of 22 degrees C and 26 degrees C at 25 degrees C and 40 degrees C, respectively. For delicate instruments, a 30-percent peroxide solution followed by an acetone rinse provides an effective alternative to classic heat sterilization.

Acetone↗