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

A template DNA from phage lambdah80dlacp5 coding for the in vitro synthesis of beta-galactosidase was used to study the effect of DNA methylation by the alkylating agent, dimethyl sulfate (DMS). Increasing the levels of DMS up to 50 mM concentration in the incubation medium led to an increase of DNA methylation. When incubated for 10 min at 37 degrees C, 3-4% Of nucleotides were methylated. The increase was linear to about 0.6% nucleotide methylation level. A higher yield was obtained at 37 degrees C incubation temperature than at 20 degrees C. Methylation of lambdah80dlacp5 DNA alone without methylation of other factors in the incubation mixture caused inhibition of the synthesis of beta-galactosidase in vitro. Increasing levels of DNA methylation caused greater inhibition of the newly synthesized enzyme activity. Total protein and RNA synthesis was inhibited by the methylated DNA to a much lesser extent than the inhibition of enzyme activity. When the level of nucleotide methylation was 0.74%, only 2% of enzyme activity remained, but total protein and RNA synthetic activities were found to be 72% and 44%, respectively.
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PMID:Effects of methylation of the beta-galactosidase genome upon in vitro synthesis of beta-galactosidase. 95 31

Methylation of a genome DNA lambda h80dlacps by N-methyl-Nitrosourea (MNU, a potent carcinogen) and dimethyl sulfate (DMS, a weak carcinogen) results in loss of its template activity in directing beta-galactosidase (beta G) synthesis in vitro, the degree of inhibition in template activity is proportional to the extent of methylation which is in turn related to the concentration of the methylating agents during 10 minutes incubation at 37 degrees C. When these methylating agents are added to the complete synthesizing system, beta G synthesis is also impaired. Maximum inhibition occurs when the chemicals are added at the initiation of this coupled transcription-translation assay. Inhibition gradually decreases at later times of addition until ten minutes after initiation when no inhibition is observed. This suggests that the early stages in mRNA synthesis are most sensitive to these agents. Preincubation of DNA with MNU for 10 min at 37 degrees C prior to the addition of other assay components (including S30 cell lysate) results in greater inhibition than preincubation of the S30 preparation with MNU prior to DNA and cofactors addition. The opposite result is obtained with DMS suggesting that while DNA is the more sensitive component of the system for MNU, S30 is the more sensitive component for DMS.
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PMID:Inhibition of DNA-directed beta-galactosidase synthesis in a cell-free system by dimethyl sulfate and N-methyl-N-nitrosourea. 679 48

Previous research has indicated that the growth rate-dependent regulation of Escherichia coli gnd expression involves the internal complementary sequence (ICS), a negative control site that lies within the 6-phosphogluconate dehydrogenase coding sequence. To determine whether the ICS acts as a transcriptional operator or attenuator, we measured beta-galactosidase-specific activities in strains carrying gnd-lac operon and protein fusions containing or lacking the ICS. Whereas the presence of the ICS repressed beta-galactosidase expression from a protein fusion by 5-fold during growth on acetate and by 2.5-fold during growth on glucose, it had no effect on beta-galactosidase expression from an operon fusion. In vitro ribosome binding experiments employing the primer extension inhibition (toeprint) assay demonstrated that the presence of the ICS in gnd mRNA reduces both the maximum extent and the rate of ternary complex formation. Moreover, the effects of deletions scanning the ICS on in vivo gene expression were highly correlated with the effects of the deletions on ribosome binding in vitro. In addition, the distal end of the ICS element was found to contribute more to ICS function than did the proximal portion, which contains the complement to the Shine-Dalgarno sequence. Finally, RNA structure mapping experiments indicated that the presence of the ICS in gnd mRNA reduces the access of the nucleotides of the ribosome binding site to the single-strand-specific chemical reagents dimethyl sulfate and kethoxal. Taken together, these data support the hypothesis that the role of the ICS in the growth rate-dependent regulation of gnd expression is to sequester the translation initiation region into a long-range mRNA secondary structure that blocks ribosome binding and thereby reduces the frequency of translation initiation.
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PMID:Inhibition of translation initiation on Escherichia coli gnd mRNA by formation of a long-range secondary structure involving the ribosome binding site and the internal complementary sequence. 759 34