Biomedical subjects

A Wipat

Publications and source records attributed to A Wipat.

8 recordsLinked to original sources

PubMed2004

SARGE: a tool for creation of putative genetic networks.

SUMMARY: SARGE is a tool for creating, visualizing and manipulating a putative genetic network from time series microarray data. The tool assigns potential edges through time-lagged correlation, incorporates a clustering mechanism, an interactive visual graph representation and employs simulated annealing for network optimization. AVAILABILITY: The application is available as a .jar file from http://www.bioinformatics.cs.ncl.ac.uk/sarge/index.html.

Algorithms↗

PubMed2003

Essential Bacillus subtilis genes.

To estimate the minimal gene set required to sustain bacterial life in nutritious conditions, we carried out a systematic inactivation of Bacillus subtilis genes. Among approximately 4,100 genes of the organism, only 192 were shown to be indispensable by this or previous work. Another 79 genes were predicted to be essential. The vast majority of essential genes were categorized in relatively few domains of cell metabolism, with about half involved in information processing, one-fifth involved in the synthesis of cell envelope and the determination of cell shape and division, and one-tenth related to cell energetics. Only 4% of essential genes encode unknown functions. Most essential genes are present throughout a wide range of Bacteria, and almost 70% can also be found in Archaea and Eucarya. However, essential genes related to cell envelope, shape, division, and respiration tend to be lost from bacteria with small genomes. Unexpectedly, most genes involved in the Embden-Meyerhof-Parnas pathway are essential. Identification of unknown and unexpected essential genes opens research avenues to better understanding of processes that sustain bacterial life.

Bacillus subtilis↗

PubMed1997

An internal FK506-binding domain is the catalytic core of the prolyl isomerase activity associated with the Bacillus subtilis trigger factor.

Two major families of peptidylprolyl cis-trans-isomerases, the cyclophilins and the structurally unrelated FK506-binding proteins (FKBPs), have been identified as cellular factors involved in protein folding in vitro. Here we report on the biochemical characterization of a second prolyl isomerase of Bacillus subtilis that was purified from a cyclophilin-negative (ppiB null) mutant and was shown to be the trigger factor (TigBS). N-terminal sequencing of 27 amino acid residues of the purified protein revealed 100% identity to the deduced sequence encoded by the tig gene, sequenced as a part of the B. subtilis genome project. The tigBS gene, located at 246 degrees on the genetic map upstream of the clpX and lonA,B genes, encodes an acidic protein (pI 4.3) of 47.5 kDa. Purified and recombinant TigBS-His proteins share the same substrate specificity and catalytic activity (Kcat/K(m) of 1.5 microM-1 s-1); both are inhibited by the macrolide FK506 with IC50 the range of 500 nM. We also demonstrate that the prolyl isomerase activity of TigBS is mediated by an internal domain of about 13 kDa (homologous to FKPB12) that represents the catalytic core of the trigger factor.

Amino Acid Isomerases↗

PubMed1996

Sequence and transcriptional analysis of clpX, a class-III heat-shock gene of Bacillus subtilis.

The nucleotide sequence of clpX, which is localized between the tig (trigger factor) and the lon (ATP-dependent protease) genes at 245 degrees on the standard Bacillus subtilis (Bs) genetic map, was determined. The putative clpX gene codes for a 46-kDa protein of 421 amino acid (aa) residues. A comparison of the deduced aa sequence with those of the recently described bacterial clpX gene products from Synechocystis sp., Escherichia coli (Ec), Haemophilus influenzae and Azotobacter vinelandii revealed strong similarities. However, in contrast to Ec, clpX and clpP of Bs are located at different loci on the chromosome and are transcribed as monocistronic genes. A heat-inducible sigma A-like promoter was mapped upstream of the clpX structural gene, but no CIRCE element, characteristic of class-I heat-shock genes (e.g., groESL and dnaK), was found between the transcriptional and translational start sites. Although the majority of the heat-inducible general stress genes in Bs are under the control of the alternative sigma factor, sigma B, the heat induction of clpX appears to be sigma B-independent. The latter indicates that clpX belongs to class-III heat-inducible genes.

ATPases Associated with Diverse Cellular Activitie↗

PubMed1996

Sequencing and functional analysis of the genome of Bacillus subtilis strain 168.

The international programme to sequence the 4.2 Mb genome of Bacillus subtilis, a model Gram-positive bacterium, is a joint project involving European, Japanese and US research groups. To date ca. 3.0 Mb of the genome has been sequenced, with the remaining 1.2 Mb expected to be completed in 1997. The amenability of B.subtilis to genetic manipulation, combined with the availability of extensive expertise on its biochemistry and physiology, makes this bacterium a valuable organism in which to investigate the properties of genes for which functions cannot be readily ascribed by standard methods.

Bacillus subtilis↗

PubMed1996

The dnaB-pheA (256 degrees-240 degrees) region of the Bacillus subtilis chromosome containing genes responsible for stress responses, the utilization of plant cell walls and primary metabolism.

Within the framework of the international programme to sequence the genome of Bacillus subtilis strain 168, we were allocated the region between dnaB (256 degrees) and pheA (240 degrees). The sequencing of this region is now complete and we report our primary analysis of the 114 kb region containing 114 ORFs. In addition to previously characterized genes, we have identified genes involved in the utilization of plant cell wall polysaccharides, stress responses and the metabolism of amino acids, cell walls, DNA and fatty acids. We also discuss various structural and physical features, including the orientation of genes with respect to replication, putative start and stop codons, ribosome binding sites and rho-independent transcription terminators.

Bacillus subtilis↗

PubMed1994

Monoclonal antibodies for Streptomyces lividans and their use for immunomagnetic capture of spores from soil.

Monoclonal antibodies were produced to Streptomyces lividans spore surface antigens. One particular hybridoma cell line, 43H6, produced a monoclonal antibody that reacted exclusively with Streptomyces cluster group 21 in an enzyme-linked immunosorbent assay (ELISA). Antibody 43H6 was found to be of subclass IgG1, kappa light chain. Western blot (immunoblot) analysis revealed that 43H6 recognized a major outer spore polypeptide of about 37,000 Da. The epitope was stably maintained in S. lividans spores over at least seven sporulation cycles on laboratory medium and for at least 14 weeks in sterile soil systems. The species group specificity of antibody 43H6 was exploited in the development of an immunocapture technique for the isolation of streptomycetes from soil. Magnetic beads coated with antibody 43H6 were mixed with soil samples seeded with S. lividans spores. Spore-bead complexes were recovered using magnets. Treatment of beads with blocking agents and the inclusion of detergents in the recovery system lessened non-specific binding of spores to beads and improved recovery. In buffer solutions decreasing the spore concentration increased the recovery values for a fixed bead concentration. At a spore concentration of 5 x 10(7) ml-1 the recovery was 4.3% whilst at 5 x 10(2) ml-1 it was 76% for a fixed bead concentration of 0.6 mg ml-1. Using a bead concentration of 2 mg per 10 g soil, approximately 30% of the target spore population of 10(6) c.f.u. was recovered from sterile soil and 4% from non-sterile soil. This method offers a rapid means of selectively recovering and concentrating Streptomyces spores from soil samples.

Antibodies, Bacterial↗

PubMed1991

Streptomyces marker plasmids for monitoring survival and spread of streptomycetes in soil.

Plasmid constructs pNW1 through pNW6 containing a controllable xylE gene (for catechol 2,3-dioxygenase) were introduced into Streptomyces lividans strains to provide a selectable marker system. xylE functions in S. lividans under the control of bacteriophage lambda promoters lambda pL and lambda pR. Thermoregulated expression of xylE is provided through the lambda repressor cI857. Catechol 2,3-dioxygenase activity was increased 2.8-fold from plasmid construct pNW2 (lambda pL, xylE, cI857) and 9.5- and 7.4-fold from constructs pNW3 (lambda pR, xylE, cI857) and pNW5 (lambda pR, xylE, cI857), respectively, when the temperature was shifted from 28 degrees C to 37 degrees C. The stability of the constructs varied from 4.7% for pNW2 to 99.4% for pNW4 (lambda pL, xylE) over two rounds of sporulation. Marked S. lividans strains released into soil systems retained the XylE phenotype for more than 80 days, depending on the marker plasmid, when examined by a selective plating method. Furthermore, S. lividans harboring plasmid pNW5 was detectable by nucleic acid hybridization at less than 10 CFU g-1 (dry weight) of soil as mycelium and 10(3) CFU g-1 (dry weight) of soil as spores with the xylE marker DNA extracted from soil and amplified by using the polymerase chain reaction.

Base Sequence↗