Biomedical subjects

Teca Calcagno Galvão

Publications and source records attributed to Teca Calcagno Galvão.

5 recordsLinked to original sources

PubMed2006

Uncoupling of choline-O-sulphate utilization from osmoprotection in Pseudomonas putida.

The genomic context of the recognized bet genes for choline-O-sulphate (COS) utilization in Pseudomonas putida KT2440 is such that betC (choline sulphatase) lies adjacent to an ATP-binding cassette transporter and a LysR type regulator, but well away from betBA, encoding enzymes for transformation of choline into glycine betaine. The consequences of such genetic layout of the functions for COS metabolism have been examined with a suite of genetic and biochemical approaches. An early clue of the utilities of the betencoded products was exposed by the phenotypes of a betC deletion. This mutant still accumulated intact COS but failed to use this compound as carbon or nitrogen source. Furthermore, betC expression was downregulated at high salt concentrations, showing that the principal role of this gene lied in COS metabolism, not in osmoprotection. In contrast, the betBA genes were required for choline transformation into the highly effective compatible solute glycine betaine (and the concomitant endurance to high salt) and also for its utilization as carbon or nitrogen source. Thus, unlike in the cases of Bacillus subtilis and Sinorhizobium meliloti, betC is unrelated to osmoprotection in Pseudomonas putida while the betBA genes are required for both betaine synthesis and tolerance to high osmotic pressure.

Bacterial Proteins↗

PubMed2005

Transcriptional regulators à la carte: engineering new effector specificities in bacterial regulatory proteins.

For many regulators of bacterial biodegradation pathways, small molecule/effector binding is the signal for triggering transcriptional activation. Thus, regulation results from a cross-talk between chemicals sensed by transcriptional factors and operon expression status. These features can be utilised in the construction of biosensors for a wide range of target compounds as, in principle, any regulatory protein whose activity is modulated by binding to a small molecule can have its effector/inducer profile artificially altered. The cognate specificities of a number of regulatory proteins have been modified as an astute approach to developing, among others, bacterial biosensors for environmentally relevant compounds.

Bacterial Proteins↗

PubMed2005

Structure-specific binding of MeCP2 to four-way junction DNA through its methyl CpG-binding domain.

MeCP2, whose methylated DNA-binding domain (MBD) binds preferentially to DNA containing 5Me-CpG relative to linear unmethylated DNA, also binds preferentially, and with similar affinity, to unmethylated four-way DNA junctions through the MBD. The Arg133Cys (R133C) mutation in the MBD, a Rett syndrome mutation that abolishes binding to methylated DNA, leads to only a slight reduction in the affinity of the MBD for four-way junctions, suggesting distinct but partially overlapping modes of binding to junction and methylated DNA. Binding to unmethylated DNA junctions is likely to involve a subset of the interactions that occur with methylated DNA. High-affinity, methylation-independent binding to four-way junctions is consistent with additional roles for MeCP2 in chromatin, beyond recognition of 5Me-CpG.

Amino Acid Sequence↗

PubMed2005

Exploring the microbial biodegradation and biotransformation gene pool.

Similar to the New World explorers of the 16th and 17th century, microbiologists today find themselves at the edge of unknown territory. It is estimated that only 0.1-1% of microorganisms can be cultivated using current techniques; the vastness of microbial lifestyles remains to be explored. Because the microbial metagenome is the largest reservoir of genes that determine enzymatic reactions, new techniques are being developed to identify the genes that underlie many valuable chemical biotransformations carried out by microbes, particularly in pathways for biodegradation of recalcitrant and xenobiotic molecules. Our knowledge of catabolic routes built on research during the past 40 years is a solid basis from which to venture on to the little-explored pathways that might exist in nature. However, it is clear that the vastness of information to be obtained requires astute experimental strategies for finding novel reactions.

Bacteria↗

PubMed2005

Adaptation of the yeast URA3 selection system to gram-negative bacteria and generation of a {delta}betCDE Pseudomonas putida strain.

A general procedure for efficient generation of gene knockouts in gram-negative bacteria by the adaptation of the Saccharomyces cerevisiae URA3 selection system is described. A Pseudomonas putida strain lacking the URA3 homolog pyrF (encoding orotidine-5'-phosphate decarboxylase) was constructed, allowing the use of a plasmid-borne copy of the gene as the target of selection. The delivery vector pTEC contains the pyrF gene and promoter, a conditional origin of replication (oriR6K), an origin of transfer (mobRK2), and an antibiotic selection marker flanked by multiple sites for cloning appropriate DNA segments. The versatility of pyrF as a selection system, allowing both positive and negative selection of the marker, and the robustness of the selection, where pyrF is associated with uracil prototrophy and fluoroorotic acid sensitivity, make this setup a powerful tool for efficient homologous gene replacement in gram-negative bacteria. The system has been instrumental for complete deletion of the P. putida choline-O-sulfate utilization operon betCDE, a mutant which could not be produced by any of the other genetic strategies available.

Bacterial Proteins↗