Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

Site-directed mutagenesis was used to identify functional domains present within the human immunodeficiency virus (HIV) tat protein. Transient cotransfection experiments showed that derivatives of tat protein with amino acid substitutions either at the amino-terminal end or at cysteine residue 22, 37, 27, or 25 were no longer able to transactivate HIV long terminal repeat-directed gene expression. Incubation of Tat expressed in Escherichia coli with zinc demonstrated that both authentic Tat and cysteine mutation derivatives could form metal-protein complexes. The tat proteins that contained alterations within the cluster of positively charged amino acid residues retained their ability to transactivate gene expression, albeit at markedly reduced levels. Indirect immunofluorescence showed that the authentic tat protein and the amino-terminal and cysteine substitution mutants all localized in the nucleus, with accumulation being most evident in the nucleolus. In contrast, nuclear accumulation was greatly reduced with the basic-substitution mutations. Consistent with this result, a fusion protein that contained amino acids GRKKR, derived from the basic region, fused to the amino-terminal end of beta-galactosidase also accumulated within the nucleus. These results demonstrate that the 14-kilodalton tat protein contains at least three distinct functional domains affecting localization and transactivation.
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PMID:Structural and functional characterization of human immunodeficiency virus tat protein. 253 18

The ACE1 gene of the yeast Saccharomyces cerevisiae is required for copper-inducible transcription of the metallothionein gene (CUP1). The sequence of the cloned ACE1 gene predicted an open reading frame for translation of a 225-amino-acid polypeptide. This polypeptide was characterized by an amino-terminal half rich in cysteine residues and positively charged amino acids. The arrangement of many of the 12 cysteines in the configuration Cys-X-Cys or Cys-X-X-Cys suggested that the ACE1 protein may bind metal ions. The carboxyl-terminal half of the ACE1 protein was devoid of cysteines but was highly acidic in nature. The ability of a bifunctional ACE1-beta-galactosidase fusion protein to accumulate in yeast cell nuclei was consistent with the possibility that ACE1 plays a direct role in the regulation of copper-inducible transcription of the yeast metallothionein gene.
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PMID:A cysteine-rich nuclear protein activates yeast metallothionein gene transcription. 265 99

The beta-galactosidase-based assay for lysine developed by Tuffnell & Payne was used to measure the bioavailabilities of cyst(e)ine, methionine, threonine and tryptophan in pronase digests of 17 foods. The digests were assayed by estimating the beta-galactosidase synthesis responses of five Escherichia coli mutants, each requiring one of the respective amino acids for protein synthesis. Deletion mutants were used whenever possible in order to ensure strain stability. Single digests of each food were assayed with 3 or 4 separate cultures of each mutant and the results were compared with those from the corresponding chemical assay. Omitting the anomalously low values for one food, the rank correlation coefficients of the bio- and chemo-assay values were 0.61 (cysteine), 0.91 (lysine), 0.95 (methionine), 0.64 (threonine) and 0.85 (tryptophan). Mean (+/- S.D.) relative amino acid bioavailabilities (casein = 100%) for the 17 foods were: cysteine, 53 +/- 23; lysine, 90 +/- 10; methionine, 95 +/- 18; threonine, 89 +/- 13; and tryptophan, 89 +/- 25. The cysteine mutant appeared to give unusually low bioavailability values except for the milk products. These amino acid mutants afford a rapid method for assaying the bioavailabilities of at least four of the five amino acids studied.
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PMID:The use of Escherichia coli mutants to measure the bioavailability of essential amino acids in foods. 265 34

Human intestinal bacteria were grown in a 3-stage continuous culture system on a medium containing complex polysaccharides and proteins as carbon and nitrogen sources. Selected bacterial populations were enumerated and glycosidase, protease and arylamidase activities measured. Comparison of arylamidase and glycosidase activities in the multichamber system (MCS) and faeces showed that the predominant faecal enzymes were also produced by bacteria growing in the MCS. After 48 d operation, porcine gastric mucin (5.8 g/d) was independently fed to vessel 1. Elevated levels of volatile fatty acid (VFA) formation showed that the glycoprotein was actively fermented. The increase in carbohydrate availability as a result of breakdown of the mucin oligosaccharides stimulated bacterial growth and activities. The enzymological measurements showed that mucin increased production of both cell-bound and extracellular glycosidases, such as beta-galactosidase, alpha-glucosidase and N-acetyl-beta-glucosaminidase. Protease activities were profoundly influenced by mucin. These were largely cell-bound in non-mucin cultures but were predominantly extracellular and collagenolytic when mucin was present. Experiments with protease inhibitors showed that cysteine proteases were the major cell-bound and extracellular enzymes in both mucin and non-mucin cultures, but that serine and metalloproteases were also present. The effect of mucin on arylamidase formation was less marked, although there was increased production of these enzymes in vessels 1 and 2 of the MCS. These results suggest that host-produced substances such as mucin glycoprotein may play a role in modulating the growth and activity of bacteria growing in the human large intestine.
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PMID:Influence of mucin on glycosidase, protease and arylamidase activities of human gut bacteria grown in a 3-stage continuous culture system. 266 79

Glycoproteins located on the luminal surface of the plasma membrane of tick gut epithelial cells, when used to vaccinate cattle, are capable of stimulating an immune response that protects cattle against subsequent tick infestation. One such tick gut glycoprotein, designated Bm86, has been purified to homogeneity and the amino acid sequences of peptide fragments generated by endoproteinase Lys-C digestion have been determined. We report here the isolation and characterization of a cDNA that encodes Bm86. The nucleotide sequence of the cDNA contains a 1982-base-pair open reading frame and predicts that Bm86 contains 650 amino acids including a 19-amino acid signal sequence and a 23-amino acid hydrophobic region adjacent to the carboxyl terminus. The main feature of the deduced protein sequence is the repeated pattern of 6 cysteine residues, suggesting the presence of several epidermal growth factor-like domains. A fusion protein consisting of 599 amino acids of Bm86 and 651 amino acids of beta-galactosidase was expressed in Escherichia coli as inclusion bodies. Ticks engorging on cattle vaccinated with these inclusion bodies were significantly damaged as a result of the immune response against the cloned antigen.
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PMID:Cloning and expression of a protective antigen from the cattle tick Boophilus microplus. 269 68

A method for rapid and effective extraction of rat liver lysosomal enzymes has been elaborated. It includes isolation of lysosomal-mitochondrial fraction by means of differential centrifugation, selective destruction of the lysosomal membrane by digitonin and centrifugal obtaining of the lysosomal matrix. Total labilization of the lysosomal membrane is achieved at 0.3 mM of the detergent. The maximal enrichment of an extract by lysosomal enzymes is observed in the range of 0.3-0.4 mM of digitonin. The level of lysosomal enzyme purification is 30.7 for cysteine cathepsins B, L, H, 24.9- for beta-galactosidase, 14.1- for acid phosphatase. The method gives high yield of lysosomal enzymes (40-80%).
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PMID:[Effective extraction of lysosomal enzymes with digitonin]. 272 13

DNA from each of two specialized transducing lambda phage, lambda dcysJIHD and lambda cysJ, has been analysed by heteroduplex mapping. The segment of the Escherichia coli chromosome carried by lambda dcysJIHD was shown to be large, approximately 18 kb in length, and to replace a large length of lambda DNA, approximately 11 kb, which includes the genes for integration and recombination. Thus lambda dcysJIHD is a bio-type transducing phage. lambda cysJ was shown to have lost very little lambda DNA and to carry about 8 kb of bacterial DNA. Sites for several restriction endonucleases were mapped in the DNA from each phage and cloning experiments located some of the genes of the cluster in relation to the restriction map. Cysteine regulation of the cloned cysJ and cysD genes was shown as well as cysteine regulation of beta-galactosidase in some constructs. The direction of transcription of the cysD gene was established, and from physical evidence the size of the 'silent section' between the cysH and cysD genes was estimated to be at least 11 kb.
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PMID:Lambda transducing phage and clones carrying genes of the cysJIHDC gene cluster of Escherichia coli K12. 296 49

Blunt-end palindromic DNA linkers with a central restriction site have been designed for the multiple reading frame insertion (abbreviated MURFI) of a sense or nonsense codon into DNA. We have utilized an amber MURFI linker, 5'CTAG TCTAGA CTAG3' to disrupt the lacZ gene, yielding truncated beta-galactosidase proteins. Conditional disruption of the tetr gene in E. coli has also been demonstrated. Nonsense codon MURFI linkers permit conditional fusion of multiple gene products while sense codon linkers can add structural elements (e.g. beta-turn, cationic segment, hydrophobic segment) or a desired amino acid to a protein (e.g. methionine, cysteine). Shotgun or alternatively site-directed insertion of the symmetric linkers is possible. The over-all length of the linker may be adjusted to retain the original reading frame, matching nucleotide additions or subtractions at recipient DNA sites. If a linker restriction site occurs elsewhere in the target DNA, single linker copies may still be inserted using non-phosphorylated linkers.
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PMID:The MURFI linker for multiple reading frame insertion of a sense or nonsense codon into DNA. 300 88

The effect of treatments with various enzymes and chemically modifying agents on [3H]muscimol binding to a purified gamma-aminobutyric acid (GABA)/benzodiazepine receptor complex from the bovine cerebral cortex was examined. Treatments with pronase, trypsin, guanidine hydrochloride, and urea significantly decreased the binding of [3H]muscimol, but dithiothreitol, N-ethylmaleimide, reduced glutathione, oxidized glutathione, cysteine, and cystine had no significant effect. These results indicate that the GABA receptor indeed consists of protein, but -SH and -S-S- groups in the protein are not involved in the exhibition of the binding activity. On the other hand, column chromatography using concanavalin A-Sepharose eluted protein having [3H]muscimol binding activity and staining of glycoprotein using an electrophoresed slab gel indicated the existence of two bands originating from the subunits of the GABA/benzodiazepine receptor complex. Furthermore, treatments with various glycosidases such as glycopeptidase A, beta-galactosidase, and alpha-mannosidase significantly increased the binding of [3H]muscimol. These results strongly suggest that GABA/benzodiazepine receptor complex is a glycoprotein and that its carbohydrate chain may be a hybrid type. Treatment with beta-galactosidase resulted in the disappearance of the low-affinity site for [3H]muscimol binding and in an increase of Bmax of the high-affinity site, without changing the KD value. These results suggest that the carbohydrate chain in the receptor complex may have a role in exhibiting the low-affinity binding site for GABA. The observation that the enhancement of [3H]muscimol binding by treatments with beta-galactosidase and glycopeptidase A were much higher than that with alpha-mannosidase may also indicate a special importance of the beta-galactosyl residue in the inhibition of GABA receptor binding activity.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Glycoprotein as a constituent of purified gamma-aminobutyric acid/benzodiazepine receptor complex: structures and physiological roles of its carbohydrate chain. 303 54

We have looked for conserved DNA sequences between four herpes simplex virus type 1 (HSV-1) glycoprotein genes encoding gB, gC, gD, and gE and pseudorabies virus (PRV) DNA, HSV-1 DNA fragments representing these four glycoprotein-coding sequences were hybridized to restriction enzyme fragments of PRV DNA by the Southern blot procedure. Specific hybridization was observed only when HSV-1 gB DNA was used as probe. This region of hybridization was localized to a 5.2-kilobase (kb) region mapping at approximately 0.15 map units on the PRV genome. Northern blot (RNA blot) analysis, with a 1.2-kb probe derived from this segment, revealed a predominant hybridizing RNA species of approximately 3 kb in PRV-infected PK15 cells. DNA sequence analysis of the region corresponding to this RNA revealed a single large open reading frame with significant nucleotide homology with the gB gene of HSV-1 KOS 321. In addition, the beginning of the sequenced PRV region also contained the end of an open reading frame with amino acid homology to HSV-1 ICP 18.5, a protein that may be involved in viral glycoprotein transport. This sequence partially overlaps the PRV gB homolog coding sequence. We have shown that the PRV gene with homology to HSV-1 gB encoded the gII glycoprotein gene by expressing a 765-base-pair segment of the PRV open reading frame in Escherichia coli as a protein fused to beta-galactosidase. Antiserum, raised in rabbits, against this fusion protein immunoprecipitated a specific family of PRV glycoproteins of apparent molecular mass 110, 68, and 55 kilodaltons that have been identified as the gII family of glycoproteins. Analysis of the predicted amino acid sequence indicated that the PRV gII protein shares 50% amino acid homology with the aligned HSV-1 gB protein. All 10 cysteine residues located outside of the signal sequence, as well as 4 of 6 potential N-linked glycosylation sites, were conserved between the two proteins. The primary protein sequence for HSV-1 gB regions known to be involved in the rate of virus entry into the cells and cell-cell fusion, as well as regions known to be associated with monoclonal antibody resistance, were highly homologous with the PRV protein sequence. Furthermore, monospecific antibody made against PRV gII immunoprecipitated HSV-1 gB from infected cells. Taken together, these findings suggest significant conservation of structure and function between the two proteins and may indicate a common evolutionary history.
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PMID:The pseudorabies virus gII gene is closely related to the gB glycoprotein gene of herpes simplex virus. 303 63


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