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 study was implemented to quantitate the hydrolase and transgalactosylase activities of beta-galactosidase (E. coli) with lactose as the substrate and to investigate various factors which affect these activities. At low lactose concentrations the rate of galactose production was equal to the rate of glucose production. The rate of galactose production relative to glucose, however, dropped dramatically at lactose concentrations higher than 0.05 M and production of trisaccharides and tetrasaccharides began (galactose/glucose ratios of about 2:1 and 3:1, respectively, were found for these two types of oligosaccharides). At least five different trissacharides were formed and their patterns of formation showed that they probably utilized both lactose and allolactose as galactosyl acceptors. Allolactose was produced in amounts proportional to glucose at all lactose concentrations (ratios of allolactose/glucose were about 0.88). Analyses of various data, including a reaction analyzed at very early times, showed that the major means of production of allolactose (and the only means initially) was the direct enzymatic transfer of galactose from the 4 position to the 6 position of the glucose moiety of lactose without prior release of glucose from the enzyme. It was shown, however, that allolactose could also be formed in significant quantities by the transfer of galactose to the 6 position of free glucose, and also by hydrolysis of preformed trisaccharide. A mechanism which fits the initial velocity data was proposed in which the steps involving the formation of an enzyme-gallactose-glucose complex, the formation and breakage of allolactose on the enzyme, and the release of glucose all seem to be of roughly equal magnitude and rate determining. Various factors affected the amounts of transgalactosylase and hydrolase activities occurring. At high pH values (greater than 7.8) the transgalactosylase/hydrolyase activity ratio increased dramatically while it decreased at low pH values (less than 6.0). At mid pH values the ratio was essentially constant. The absence of Mg2+ caused a large decrease in the transgalactosylase/hydrolase activity ratio while the absence of all but traces of Na+ or K+ had no effect. The anomeric configuration of lactose altered the transgalactosylase/hydrolase activity ratios, alpha-Lactose resulted in a decrease of allolactose production (transgalactosylase activity) relative to hydrolase activities (glucose production) while beta-lactose had the opposite effect.
 ...
PMID:A quantitation of the factors which affect the hydrolase and transgalactosylase activities of beta-galactosidase (E. coli) on lactose. 0 22

An isogenic pair of Escherichia coli strains, one carrying an rnc+ and the other an rnc- allele (a mutation which reduces the level of ribonuclease III), was compared. The rnc- strain fails to grow at very elevated temperatures (for E. coli) while the rnc+ strain does grow exponentially. Assaying the residual RNase III like activity in extracts of the rnc- strain at different pHs and at different temperatures suggested that this residual RNase III like activity is not due to RNase III. This raised the possibility that the rnc- strain is devoid of any RNase III activity in the cell. Comparing the decay of newly synthesized RNA and functional decay of beta-galactosidase mRNA in such strains revealed that in both strains these parameters proceed in similar rates, which suggests that RNase III is not involved in the metabolism of mRNA. During carbon starvation preexisting total RNA, as well as 23S and 16S rRNA, decay faster in the rnc- strain, thus eliminating the possibility that RNase III is the endoribonuclease which initiates the decay of rRNA during starvation (Kaplan and Apirion, 1975a).
 ...
PMID:Consequences of losing ribonuclease III on the Escherichia coli cell. 77 91

Immunoreactivities of recombinant carcinoembryonic antigen (CEA) proteins expressed in Escherichia coli (E. coli) were analyzed in relation to the CEA domain structure [domains N, I (A1-B1), II (A2-B2), III (A3-B3) and M]. We reconstructed in a prokaryotic expression vector, pUCPL-cI, the cDNAs fro CEA-N, CEA-I, CEA-II, and CEA-III-M. The latter three were expressed as fusion products with bacterial beta-galactosidase. The recombinant proteins were solubilized by sonication in 1% sodium dodecyl sulfate (SDS) and purified by preparative SDS-polyacrylamide gel electrophoresis followed by electroelution. Their molecular weights judged from Western blotting coincided with those calculated from their cDNA sequences, respectively. By solid-phase enzyme immunoassays, the immunoreactivities of the purified recombinant proteins were tested with 21 distinct anti-CEA monoclonal antibodies (MAbs) which had been found to recognize the peptide epitopes of the CEA molecule and to be reactive with the recombinant CEA proteins expressed in Chinese hamster ovary (CHO) cells. Fourteen of the 21 MAbs reacted with the recombinant CEA proteins expressed in E. coli and confirmed the localization of the epitopes identified by using the recombinant CEA proteins expressed in CHO cells. The reactivities of 5 MAbs with the recombinant proteins expressed in E. coli were remarkably low when compared with those of the proteins expressed in CHO cells but also confirmed the localization of the epitopes identified with the recombinant CEA proteins expressed in CHO cells. The remaining 2 MAbs did not react with any recombinant protein expressed in E. coli. These results indicate that the fusion CEA-proteins expressed in E. coli are useful in the localization of the epitopes on the polypeptide chains when they reacted with the MAbs tested. However, one third of the epitopes of CEA peptides may be profoundly affected by the presence of disulfide bonds and/or sugar chains which do not seem to be formed well in E. coli.
 ...
PMID:Immunoreactivity of recombinant carcinoembryonic antigen proteins expressed in Escherichia coli. 137 88

Glutamic acid 461 of beta-galactosidase (E. coli) was replaced by gln using site-directed mutagenesis. Kinetic studies on the purified Q461-beta-galactosidase showed that it had less than 0.4% of the wild-type activity (with ONPG as substrate), confirming other studies which have suggested that the negative charge on glu-461 is important for activity. The Km values did not increase, indicating that binding of the substrate was not decreased by this change. Thermal denaturation studies showed Q461-beta-galactosidase to be somewhat more susceptible to heat denaturation than the wild-type enzyme.
 ...
PMID:Site-directed mutagenic replacement of glu-461 with gln in beta-galactosidase (E. coli): evidence that glu-461 is important for activity. 289 51

This paper describes a new sensitive microELISA based on enzyme/anti-enzyme complexes following an unlabelled antibody bridging step. beta-Galactosidase/anti-beta-galactosidase complexes were made using a monoclonal antibody raised against bacterial (E. coli) beta-galactosidase and enzyme activity was quantified with a fluorogenic substrate. Because of its high sensitivity the assay is particularly suitable for the detection of limited amounts of antigen. One application illustrated is the analysis of Class I and Class II histocompatibility antigens on peripheral blood lymphocytes using 5000 cells/well in 60-well Terasaki or 96-well microtitre plates.
 ...
PMID:A sensitive micro-immunoassay using beta-galactosidase/anti-beta-galactosidase complexes. 310 11

By using the technique of site-directed mutagenesis we have succeeded in replacing tyr-503 of beta-galactosidase (E. coli) with a phe. A study of the kinetic and stability properties of this mutant enzyme (F-503 beta-galactosidase) showed that the loss in activity upon this change is due to the loss of a catalytic group (rather than a detrimental change in the enzyme's overall structure or a change in the enzyme's binding capacity). This confirms previous suggestions that this tyr residue is involved in catalysis.
 ...
PMID:Site-directed mutagenesis of beta-galactosidase (E. coli) reveals that tyr-503 is essential for activity. 313 51

Glu-416 of beta-galactosidase (E. coli) was replaced with Gln and Val using site-directed mutagenesis. The substituted enzymes displayed a greatly decreased sensitivity to Mg2+. Equilibrium dialysis studies indicated that wild-type beta-galactosidase bound Mg2+ tightly, whereas E416V-beta-galactosidase did not. In addition, the pH profile of E416V-beta-galactosidase was unaffected by the presence or absence of 1 mM Mg2+. Surprisingly, both substituted enzymes were inactivated, rather than activated, by Mg2+ but high amounts of Mg2+ were needed (1 mM). E416Q-beta-galactosidase was unstable when stored in the presence of Mg2+. The substituted enzymes displayed a dramatically lowered affinity for the synthetic substrate, ONPG, and for IPTG (a substrate analog inhibitor) in both the presence and the absence of Mg2+.
 ...
PMID:Glu-416 of beta-galactosidase (Escherichia coli) is a Mg2+ ligand and beta-galactosidases with substitutions for Glu-416 are inactivated, rather than activated, by MG2+. 861 91

The activation of beta-galactosidase (E. coli) by Mg(2+) at pH values below 7.6 was studied. If the Mg(2+) concentration was high enough, the k(cat) values at pH values down to 5.0 remained at the same high level as at pH 7 and 7.6 (600-620 s-(1) with o-nitrophenyl-beta-D-galactopyranoside as the substrate). Very high concentrations of Mg(2+) (greater than 100 mM at pH 5) were, however, needed to saturate the Mg(2+) site at lower pH values. The Km values at low levels of Mg(2+) were high at every pH but they decreased and approached the same low value at every pH (about 0.13 mM) as the [Mg(2+)] was increased. These data indicate that it is difficult to bind Mg(2+) at lower pH values, but the k(cat) and K(m) values of the enzyme, and therefore the rates of galactosylation (k(2)), degalactosylation (k(3)), and binding (K(s)), do not change substantially as a function of pH provided that a Mg(2+) is bound to the enzyme. The data also showed that Mg(2+) and protons compete for the same site. Analysis by plotting log [Mg(2+)](mid) vs. pH showed that the binding of Mg(2+) to the free enzyme involves two groups with pK(a) values in the vicinity of 7 and one group with a pK(a) value near 5.5. (The values referred to as [Mg(2+)](mid) are the Mg(2+) concentrations that resulted in k(cat) values midway between basal and maximum.) The "apparent" pK(a) values of the groups when a Mg(2+) was bound (at saturating [Mg(2+)]) all appeared to be below 5.0.
 ...
PMID:The activation of beta-galactosidase (E. coli) by Mg(2+) at lower pH values. 921 40

Somatic intrachromosomal recombination (SICR) can result in inversions and deletions in the DNA. pKZ1 mice possess an Escherichia coli (E. coli) lacZ transgene which is only expressed after a DNA inversion involving the transgene occurs. The E. coli beta-galactosidase protein can then be detected in frozen tissue sections using a chromogenic substrate. Therefore, pKZ1 mice can be used to detect SICR inversion events in vivo in different tissues. We have tested the pKZ1 mouse for its potential as a general mutagenesis model for detecting SICR in spleen in response to carcinogens which have widely different mechanisms of genotoxicity. Animals were given a single exposure of carcinogen and spleen cells were examined 3 days later for inversion events by histochemical staining of tissue sections. Mitomycin C, X-irradiation, etoposide and methylene chloride caused significant induction of inversion events in spleen tissue, ranging from 1.6- to 4.2-fold induction with the doses used here. This is the first time that inversion events induced by these carcinogens have been specifically studied in vivo in a mouse model and the findings expand the repertoire of mutation events known to be caused by these agents. We suggest that the pKZ1 mouse can be used as a general mutagenesis model for detection of SICR events and is likely to be a useful model for studying the mechanism of SICR in response to DNA damaging agents.
 ...
PMID:Inversion due to intrachromosomal recombination produced by carcinogens in a transgenic mouse model. 1035 96

This study compared the performance of LMX(R) broth (LMX), Chromocult Coliform(R) agar (CC) and Chromocult Coliform agar plus cefsulodin (10 microg ml-1) (CC-CFS), with standard methods multiple tube fermentation (MTF), for the enumeration of total coliforms and Escherichia coli from marine recreational waters. LMX and CC are two media designed to concurrently detect total coliform (TC) bacteria and E. coli by the specific action of beta-galactosidase (total coliforms) and beta-glucuronidase (E. coli). Overall results for the TC test showed that LMX, CC and MTF recovered 2.63, 1.95 and 1.90 times as many TCs as CC-CFS, respectively. Data from the multiple range test showed significant differences (P < 0.05) between TC counts on CC-CFS and LMX. The traditional MTF was less sensitive for E. coli enumeration. However, there was no statistically significant differences between LMX, CC, CC-CFS and the MTF method for E. coli enumeration. Background interference was reduced on CC-CFS and the counts obtained reflected more accurately the number of TCs. Therefore, the contribution of beta-galactosidase positive, non coliform bacteria (Aeromonas spp. and Vibrio spp.) to TC counts should not be neglected.
 ...
PMID:Quantitative determination of total coliforms and Escherichia coli in marine waters with chromogenic and fluorogenic media. 1073 96


1 2 3 Next >>