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)

TraJ and SfrA are, respectively, plasmid and host (Escherichia coli)-encoded proteins normally required for F plasmid traY promoter function. Beginning with plasmids in which a traY-lacZ fusion gene, designated phi (traY'-'lacZ)hyb, and lacY are expressed from the F plasmid traY promoter, we isolated mutants in which lac gene expression was SfrA or TraJ-independent. A total of 45 of 50 SfrA-independent isolates obtained after 2-aminopurine mutagenesis proved to have chromosomal mutations, whereas four out of four isolates obtained without mutagenesis had plasmid mutations. All of 17 isolates selected for TraJ-independent expression after mutagenesis had plasmid mutations. By restriction endonuclease digestions, 25 of 26 SfrA-independent and TraJ-independent plasmid mutations were insertions. Four of the former and three of the latter were examined further. By sequence analysis, all seven proved to be IS1 or IS2 insertions defining five insertion sites between base-pairs -49 and -82 with respect to the major traY transcription initiation site. In two cases, the same insertion allele was obtained from the two selection schemes. All three of the mutants selected for TraJ-independent gene expression manifested SfrA-independent expression as well, and levels of beta-galactosidase in different plasmid mutant strains lacking TraJ and SfrA were indistinguishable. By primer extension analysis, transcription initiation sites for traY mRNA synthesis were unaltered by the mutations. Replacing the tra sequence upstream from base-pair -78, without genetic selection, increased beta-galactosidase activity in the absence of TraJ and SfrA greater than tenfold. Activity increased two- to threefold more in a traJ+ sfrA mutant strain, and fivefold more in a traJ+ sfrA+ strain. Activity was unaltered in an sfrA+ strain without TraJ. By primer extension analysis, the traY promoter was utilized under all conditions. The data indicate that regulation of traY promoter activity is strongly dependent on sequence context.
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PMID:Regulation of the F plasmid traY promoter in Escherichia coli K12 as a function of sequence context. 190 41

The IS2 sequence encodes five open reading frames (ORF1 to ORF5) that are greater than 150 nucleotides each. Only one protein of 14 kDa was detected when the expression of IS2 genes was examined in minicells. This 14 kDa protein was referred to as InsA in this study and was determined to be encoded by ORF1. A sixfold decrease in IS2 transposition frequency was observed when insA was overexpressed. DNA footprinting results indicated that InsA binds to the sequence 5'-TAAATAA-3' located at IS2 nucleotide numbers 1286 to 1292. (The IS2 right terminal repeat spans nucleotides 1290 to 1331.) This InsA binding sequence is situated 4 bp upstream from the putative "-10" sequence of the insA promoter that overlaps the right terminal repeat of IS2. The presence of a promoter located in this region was demonstrated by the ability of a DNA fragment containing the right terminal repeat to drive the expression of a promoterless lacZ gene. The transcription of insA was determined to start at the A residue located at nucleotide number 1268. With the same insA promoter-lacZ fusion construct, overexpression of insA in the same cell was found to decrease the beta-galactosidase activity. The results of this study suggest that InsA affects IS2 transposition by regulating the transcription of IS2 genes.
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PMID:Functional analysis of the 14 kDa protein of insertion sequence 2. 810 36

The biosynthesis of Mn-containing superoxide dismutase is regulated in response to stimuli that affect the redox potential of the cell. To further investigate the mode of regulation of the gene (sodA) encoding this enzyme, cis-acting regulatory mutations in a strain containing a sodA::lacZ gene fusion were studied. The mutant strains expressed beta-galactosidase under anaerobic conditions, whereas the wild-type did not. Furthermore, the mutants were not induced in response to the presence of iron chelator, 2,2'-dipyridyl, or to the redox cycling compound, paraquat. The wild-type, however, did respond to these effectors. In vivo cloning was used to isolate the cis-acting regulatory elements from the mutants (NC4 and NC5). Replacement of the wild-type 5'-regulatory region with either of the mutants' cis-acting regulatory element resulted in the anaerobic expression of active Mn-superoxide dismutase. Sequence and restriction analysis revealed the presence of an IS2 insertion element in the promoter region of one of the mutants (NC5). This insertion caused the displacement of the 5'-regulatory region of sodA and the formation of a functional hybrid promoter consisting of the resident-10 region from sodA and -35 from IS2. The second mutation (from NC4) was similarly analyzed, and an IS5 element was identified. The insertion site of IS5 (in NC4) was 6 bp (5'-TTAATT-3') upstream from the IS2 site (in NC5). Anaerobic expression of sodA in NC4 was lower than in NC5. This difference was almost eliminated in an arc- background, suggesting that the sequence 5'-TTAATT-3' might be essential for negative regulation by ArcA.
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PMID:Characterization of cis-acting regulatory mutations causing anaerobic expression of the sodA gene in Escherichia coli. 838 43

p94/calpain 3 is a skeletal muscle-specific member of the Ca(2+)-regulated cytosolic cysteine protease family, the calpains. Defective p94 protease activity originating from gene mutations causes a muscular dystrophy called calpainopathy, indicating the indispensability of p94 for muscle survival. Because of the existence of the p94-specific regions IS1 and IS2, p94 undergoes very rapid and exhaustive autolysis. To elucidate the physiological relevance of this unique activity, the autolytic profiles of p94 and the effect of the p94 binding protein, connectin/titin, on this process were investigated. In vitro analysis of p94 autolysis showed that autolysis in IS1 proceeds without immediate disassembly into fragments and that the newly identified cryptic autolytic site in IS2 is critical for disassembling autolyzed fragments. As a genetic system to assay p94 autolysis semiquantitatively, p94 was expressed in yeast as a hybrid protein between the DNA binding and activation domains of the yeast transcriptional activator Gal4. Transcriptional activation by the Gal4-p94:WT hybrid protein is precluded by p94 autolysis. Complete or partial loss of autolytic activity by C129S active site mutation, limb girdle muscular dystrophy type 2A pathogenic missense mutations, or PCR-based random mutagenesis could be detected by semiquantitative restoration of Gal4-dependent beta-galactosidase gene expression. Using this system, the N2A connectin fragment that binds to p94 was shown to suppress p94 autolytic disassembly. The proximity of the IS2 autolytic and connectin-binding sites in p94 suggested that N2A connectin suppresses IS2 autolysis. These data indicate the importance of p94-connectin interaction in the control of p94 functions by regulating autolytic decay of p94.
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PMID:Suppressed disassembly of autolyzing p94/CAPN3 by N2A connectin/titin in a genetic reporter system. 1662 76