Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.2.1.23 (beta-galactosidase)
 14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A recessive mutation, aarG1, has been identified that resulted in an 18-fold increase in the expression of beta-galactosidase from an aac(2')-lacZ fusion. Transcriptional fusions and Northern blot analysis demonstrated that the aarG1 allele also resulted in a large increase in the expression of aarP, a gene encoding a transcriptional activator of aac(2')-Ia. The effects of aarG1 on aac(2')-Ia expression were mediated by aarP-dependent and -independent mechanisms. The aarG1 allele also resulted in a multiple antibiotic resistance (Mar) phenotype, which included increased chloramphenicol, tetracycline and fluoroquinolone resistance. This Mar phenotype also resulted from aarP-dependent and -independent mechanisms. Sequence analysis of the aarG locus revealed the presence of two open reading frames, designated aarR and aarG, organized in tandem. The putative AarR protein displayed 75% amino acid identity to the response regulator PhoP, and the AarG protein displayed 57% amino acid identity to the sensor kinase PhoQ. The aarG1 mutation, a C to T substitution, resulted in a threonine to isoleucine substitution at position 279 (T279I) in the putative sensor kinase. The AarG product was functionally similar to PhoQ, as it was able to restore wild-type levels of maganin resistance to a Salmonella typhimurium phoQ mutant. However, expression of the aarP and aac(2')-Ia genes was not significantly affected by the levels of Mg2+ or Ca2+, suggesting that aarG senses a signal other than divalent cations.
 ...
PMID:A regulatory cascade involving AarG, a putative sensor kinase, controls the expression of the 2'-N-acetyltransferase and an intrinsic multiple antibiotic resistance (Mar) response in Providencia stuartii. 968 Feb 22

Hydropathy profile analysis of the amino acid sequence of the Na+/proline transporter of Escherichia coli (PutP) suggests that the protein consists of 12 transmembrane domains (TMs) which are connected by hydrophilic loops (Nakao, T., Yamato, I., and Anraku, Y. (1987) Mol. Gen. Genet. 208, 70-75). We have tested this prediction by applying a gene fusion approach in combination with a Cys accessibility analysis and site-specific proteolysis. Characterization of a series of PutP-alkaline phosphatase (PhoA) and PutP-beta-galactosidase (LacZ) hybrid proteins yields a reciprocal activity pattern of the reporter proteins that is in agreement with the topology of TMs III to XII of the 12-helix model. Placement of the PutP-PhoA and PutP-LacZ junction sites closer to the N terminus does not yield conclusive results. As a prerequisite for further topology studies, a functional PutP molecule devoid of all five native Cys residues (Cys-free PutP) is generated. Subsequently, amino acids in Cys-free PutP are replaced individually with Cys, and the accessibility of the sulfhydryl groups is analyzed. Surprisingly, Cys residues placed close to the N terminus of PutP (Ile-3 --> Cys, Thr-5 --> Cys) or into putative TM II (Ser-71 --> Cys, Glu-75 --> Cys) are highly accessible to membrane permeant and impermeant thiol reagents in intact cells. In contrast, Cys at the C terminus (Ser-502 --> Cys) reacts only with the membrane permeant but not with the impermeant reagent in intact cells. These results contradict the 12-helix motif and indicate a periplasmic location of the N terminus whereas the C terminus faces the cytoplasm. In addition, a transporter with Cys in place of Leu-37 (putative periplasmic loop (pL2) shows the same accessibility pattern as the Cys at the C terminus. Furthermore, PutP which has been purified and reconstituted into proteoliposomes in an inside-out orientation, is readily cleaved by the endoproteinase AspN before Asp-33 (pL2), Asp-112 (putative cytoplasmic loop (cL3), Asp-262 (cL7), and Asp-356 (cL9). These results suggest a cytosolic location of Asp-33 and Leu-37, thereby implying the formation of an additional TM formed by amino acids of pL2. Based on these observations, a new secondary structure model is proposed according to which the protein consists of 13 TMs with the N terminus on the outside and the C terminus facing the cytoplasm. The 13-helix structure is discussed as a common topological motif for all members of the Na+/solute cotransporter family.
 ...
PMID:Topology of the Na+/proline transporter of Escherichia coli. 975 72

The N-end rule relates the in vivo half-life of a protein to the identity of its N-terminal residue. Ubr1p, the recognition (E3) component of the Saccharomyces cerevisiae N-end rule pathway, contains at least two substrate-binding sites. The type 1 site is specific for N-terminal basic residues Arg, Lys, and His. The type 2 site is specific for N-terminal bulky hydrophobic residues Phe, Leu, Trp, Tyr, and Ile. Previous work has shown that dipeptides bearing either type 1 or type 2 N-terminal residues act as weak but specific inhibitors of the N-end rule pathway. We took advantage of the two-site architecture of Ubr1p to explore the feasibility of bivalent N-end rule inhibitors, whose expected higher efficacy would result from higher affinity of the cooperative (bivalent) binding to Ubr1p. The inhibitor comprised mixed tetramers of beta-galactosidase that bore both N-terminal Arg (type 1 residue) and N-terminal Leu (type 2 residue) but that were resistant to proteolysis in vivo. Expression of these constructs in S. cerevisiae inhibited the N-end rule pathway much more strongly than the expression of otherwise identical beta-galactosidase tetramers whose N-terminal residues were exclusively Arg or exclusively Leu. In addition to demonstrating spatial proximity between the type 1 and type 2 substrate-binding sites of Ubr1p, these results provide a route to high affinity inhibitors of the N-end rule pathway.
 ...
PMID:Bivalent inhibitor of the N-end rule pathway. 1036 69

We increased drastically the heat stability of Lac repressor (LacR) of Escherichia coli. Wild-type tetrameric LacR denatures irreversibly at 53 degrees C. Improving hydrophobic packing at the dimerisation interface by a single substitution increases LacR heat-resistance by 40 deg. C without abolishing inducer binding at high and low temperatures. Tetrameric LacR mutants carrying substitutions of the positively charged amino acid Lys84 by each of the hydrophobic amino acids Leu, Ile and Met resist heating to temperatures up to 93 degrees C. We performed IPTG binding assays at 80 degrees C and found the mutant Lac repressors active and, thus, the core intact. Furthermore, the activity of LacR following heating is shown at room temperature by a gel retardation assay, which demonstrates normal oligomerisation state and function of the headpiece. The same mutations (K84L/I/M) in the dimer LacR331stop, carrying a stop codon in amino acid 331, increase thermostability of the dimer from 47 degrees C to 87 degrees C. LacRK84M represses beta-galactosidase activity in vivo as well as the wild-type and is sufficiently induced to allow growth on lactose. The results with both tetramer and dimer variants of LacR indicate mutual stabilisation of the tetramerisation region and the stable core.
 ...
PMID:Strengthening the dimerisation interface of Lac repressor increases its thermostability by 40 deg. C. 1083 85

Pseudomonas fluorescens F113 produces antifungal metabolites that protect the roots of sugarbeet from the fungus Pythium ultimum. The phytopathogen, in turn, has the ability to downregulate the expression of genes fundamental to the rhizosphere competence of the bacterial strain. This paper describes the characterization of two of these genes, which were isolated by screening a mini-Tn5::lacZ mutant bank for differential expression of beta-galactosidase in the presence of P. ultimum. In order to identify the genes affected in reporter mutants SF3 and SF5, the transposons and flanking regions were cloned. Sequence analysis of the regions flanking the transposons in SF3 revealed that mini-Tn5::lacZ had inserted into a tRNA(Ile) gene, which maps within a ribosomal RNA (rrn) operon. In SF5, the transposon inserted between the promoter of a second rrn operon and a gene encoding a 16S rRNA. Southern blot analysis demonstrated that there are five rrn operons in P. fluorescens F113 and that the transposons in SF3 and SF5 had inserted into two different operons. Further characterization of these mutants suggests that their reduced rhizosphere competence is not the result of reduced viability in the short term but may be accounted for partly by reduced growth rates under conditions that support rapid growth. Analysis of lacZ expression in the reporter mutants indicate that the marked rrn operons are regulated differently, suggesting different physiological roles.
 ...
PMID:Signalling by the fungus Pythium ultimum represses expression of two ribosomal RNA operons with key roles in the rhizosphere ecology of Pseudomonas fluorescens F113. 1120 71

A cDNA coding thioredoxin (TRX) was isolated from a cDNA library of Schizosaccharomyces pombe by colony hybridization. The 438 bp EcoRI fragment, which was detected by Southern hybridization, reveals an open reading frame which encodes a protein of 103 amino acids. The genomic DNA encoding TRX was also isolated from S. pombe chromosomal DNA using PCR. The cloned sequence contains 1795 bp and encodes a protein of 103 amino acids. However, the C-terminal region obtained from the cDNA clone is -Val-Arg-Leu-Asn-Arg-Ser-Leu, whereas the C-terminal region deduced from the genomic DNA appears to contain -Ala-Ser-Ile-Lys-Ala-Asn-Leu. This indicates that S. pombe cells contain two kinds of TRX genes which have dissimilar amino acid sequences only at the C-terminal regions. The heterologous TRX 1C produced from the cDNA clone could be used as a subunit of T7 DNA polymerase, while the TRX 1G from the genomic DNA could not. The upstream sequence and the region encoding the N-terminal 18 amino acids of the genomic DNA were fused into the promoterless beta-galactosidase gene of the shuttle vector YEp357 to generate the fusion plasmid pYKT24. Synthesis of beta-galactosidase from the fusion plasmid was found to be enhanced by hydrogen peroxide, menadione and aluminum chloride. It indicates that the expression of the cloned TRX gene is induced by oxidative stress.
 ...
PMID:Characterization and regulation of Schizosaccharomyces pombe gene encoding thioredoxin. 1126 79

Staphylococcus xylosus is a facultative anaerobic bacterium used as a starter culture for fermented meat products. In an attempt to analyze the antioxidant capacities of this organism, the superoxide dismutase (SOD) was characterized. S. xylosus contains a single cytoplasmic SOD, which was not inhibited by H2O2. The SOD activity in crude extracts was completely lost upon metal depletion, but it could be recovered by manganese and very weakly by iron. It is therefore suggested that the S. xylosus SOD is a manganese-preferring enzyme. The corresponding gene, sod, was isolated from a genomic library of S. xylosus DNA and complemented the growth defect of an Escherichia coli SOD-deficient mutant. As deduced from the nucleotide sequence, sod encodes a protein of 199 amino acids with a molecular mass of 22.5 kDa. Two transcriptional start sites 25 and 120 bp upstream of the sod start codon were identified. A terminator-like structure downstream of the gene suggested a monocistronic sod mRNA. Regulation of sod expression was studied using fusions of the sod promoters to a genomic promoterless beta-galactosidase gene. The sod expression was not affected by manganese and increased slightly with paraquat. It was induced during stationary phase in a complex medium but not in a chemically defined medium. To investigate the physiological role of SOD, a mutant devoid of SOD activity was constructed. Growth experiments showed that sod is not essential for aerobic growth in complex medium. However, in chemically defined medium without leucine, isoleucine, and valine, the sod mutant hardly grew, in contrast to the wild-type strain. In addition, the mutant was sensitive to hyperbaric oxygen and to paraquat. Therefore, sod plays an important role in the protection of S. xylosus from oxidative stress.
 ...
PMID:Characterization of the single superoxide dismutase of Staphylococcus xylosus. 1152 11

The activity of the promoter regions of the cell division genes ftsZ, ftsE, minC, minD and minE from Neisseria gonorrhoeae (Ng) was studied under different environmental conditions using lacZ translational fusions. The promoters of the minNg genes have not been previously determined and we identified promoter regions upstream of each gene (minCp, minDp and minEp). We determined that minDp had the strongest activity. Expression of the promoter regions of ftSZ(Ng) and ftsE(Ng), which we had previously identified, as well as minD(Ng), were then studied under conditions reflecting the environment of the genitourinary tract. These conditions included anaerobiosis, presence of isoleucine or urea (3 mM and 400 mM, respectively) and acidity of pH 6. Both beta-galactosidase expression and northern blot analysis indicated that all three genes were upregulated under anaerobiosis. The addition of isoleucine as well as media at pH 6 did not have any significant effects on the promoter activity of these genes while the presence of urea significantly decreased ftsZ(Ng) promoter activity. The expression of the minD(Ng) promoter region was analyzed during different growth phases and shown to follow the growth behavior of the culture. By contrast, the ftSZ(Ng) promoter activity continued to rise after the onset of the stationary phase. When gonococcal ftsZ promoter 1, (Pz1) was altered by site-directed mutagenesis, a significant decrease in the expression of ftsZ(Ng) was observed under both aerobic and anaerobic conditions. These data infer that gonococci regulate their cell division in response to different environments.
 ...
PMID:Expression of Neisseria gonorrhoeae cell division genes ftsZ, ftsE and minD is influenced by environmental conditions. 1176 38

A mouse bearing a novel transgene encoding the human VPAC2 receptor (hVIPR; Shen et al. (2000) PNAS, 97, 11575-11580) was used to investigate circadian function in the hypothalamic suprachiasmatic nuclei (SCN). Neurons expressing hVPAC2R, detected by a beta-galactosidase (beta-GAL) tag, have a distinct distribution within the SCN, closely matching that of neurophysin (NP) neurons and extending into the region of peptide histidine isoleucine (PHI) cells. In common with NP and PHI cells, neurons expressing hVPAC2R are circadian in nature, as revealed by synchronous rhythmic expression of mPERIOD (mPER) proteins. A population of SCN cells not expressing PHI, NP or hVPAC2R exhibited circadian PER expression antiphasic with the rest of the SCN. Nocturnal light exposure induced mPER1 in the ventral SCN and mPER2 widely across the nucleus. Induction of nuclear mPER2 in hVPAC2R cells confirmed their photic responsiveness. Having established their circadian properties, we tested the utility of SCN neurons expressing the hVIPR transgene as functionally and anatomically explicit markers for SCN tissue grafts. Prenatal SCN tissue from hVIPR transgenic pups survived transplantation into adult CD1 mice, and expressed beta-GAL, PER and PHI. Over a series of studies, hVIPR transgenic SCN grafts restored circadian activity rhythms to 17 of 72 arrhythmic SCN lesioned recipients (23.6%). By using heterozygous hVIPR transgenic grafts on a heterozygous Clock mutant background we confirmed that restored activity rhythms were conferred by the donor tissue. We conclude that the hVIPR transgene is a powerful and flexible tool for examination of circadian function in the mouse SCN.
 ...
PMID:A hVIPR transgene as a novel tool for the analysis of circadian function in the mouse suprachiasmatic nucleus. 1281 56

A mouse bearing a novel transgene encoding the human VPAC2 receptor (hVIPR; Shen et al. (2000) PNAS, 97, 11575-11580) was used to investigate circadian function in the hypothalamic suprachiasmatic nuclei (SCN). Neurons expressing hVPAC2R, detected by a beta-galactosidase (beta-GAL) tag, have a distinct distribution within the SCN, closely matching that of neurophysin (NP) neurons and extending into the region of peptide histidine isoleucine (PHI) cells. In common with NP and PHI cells, neurons expressing hVPAC2R are circadian in nature, as revealed by synchronous rhythmic expression of mPERIOD (mPER) proteins. A population of SCN cells not expressing PHI, NP or hVPAC2R exhibited circadian PER expression antiphasic with the rest of the SCN. Nocturnal light exposure induced mPER1 in the ventral SCN and mPER2 widely across the nucleus. Induction of nuclear mPER2 in hVPAC2R cells confirmed their photic responsiveness. Having established their circadian properties, we tested the utility of SCN neurons expressing the hVIPR transgene as functionally and anatomically explicit markers for SCN tissue grafts. Prenatal SCN tissue from hVIPR transgenic pups survived transplantation into adult CD1 mice, and expressed beta-GAL, PER and PHI. Over a series of studies, hVIPR transgenic SCN grafts restored circadian activity rhythms to 17 of 72 arrhythmic SCN lesioned recipients (23.6%). By using heterozygous hVIPR transgenic grafts on a heterozygous Clock mutant background we confirmed that restored activity rhythms were conferred by the donor tissue. We conclude that the hVIPR transgene is a powerful and flexible tool for examination of circadian function in the mouse SCN.
 ...
PMID:A hVIPR transgene as a novel tool for the analysis of circadian function in the mouse suprachiasmatic nucleus. 1260 72


<< Previous 1 2 3 4 5 6 Next >>