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Query: EC:3.2.1.26 (invertase)
 4,927 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Saccharomyces cerevisiae mutants containing different point mutations in the HXK2 gene were used to study the relationship between phosphorylation by hexokinase II and glucose repression in yeast cells. Mutants showing different levels of hexokinase activity were examined for the degree of glucose repression as indicated by the levels of invertase activity. The levels of hexokinase activity and invertase activity showed a strong inverse correlation, with a few exceptions attributable to very unstable hexokinase II proteins. The in vivo hexokinase II activity was determined by measuring growth rates, using fructose as a carbon source. This in vivo hexokinase II activity was similarly inversely correlated with invertase activity. Several hxk2 alleles were transferred to multicopy plasmids to study the effects of increasing the amounts of mutant proteins. The cells that contained the multicopy plasmids exhibited less invertase and more hexokinase activity, further strengthening the correlation. These results strongly support the hypothesis that the phosphorylation activity of hexokinase II is correlated with glucose repression.
Mol Cell Biol 1989 Dec
PMID:The residual enzymatic phosphorylation activity of hexokinase II mutants is correlated with glucose repression in Saccharomyces cerevisiae. 268 72

Several hundred new mutations in the gene (HXK2) encoding hexokinase II of Saccharomyces cerevisiae were isolated, and a subset of them was mapped, resulting in a fine-structure genetic map. Among the mutations that were sequenced, 35 were independent missense mutations. The mutations were obtained by mutagenesis of cloned HXK2 DNA carried on a low-copy-number plasmid vector and screened for a number of different phenotypes in yeast strains bearing chromosomal hxk1 and hxk2 null mutations. Some of these mutants were characterized both in vivo and in vitro; they displayed a wide spectrum of residual hexokinase activities, as indicated by three assays: in vitro enzyme activity, ability to grow on glucose and fructose, and ability to repress invertase production when growing on glucose. Of those that failed to support growth on fructose, only a small minority made normal-size, stable, and inactive protein. Analysis of the amino acid changes in these mutants in light of the crystallographically determined three-dimensional structure of hexokinase II suggests important roles in structure or catalysis for six amino acid residues, only two of which are near the active site.
Mol Cell Biol 1989 Dec
PMID:Isolation and characterization of mutations in the HXK2 gene of Saccharomyces cerevisiae. 268 71

A plasmid vector, pYZ1, was constructed which lacks most of the beta-lactamase signal-peptide coding region, but has a unique EcoRI site spanning codons 2 and 3 of the resultant cytoplasmic beta-lactamase derivative. Short quasi-random DNA sequences were cloned into the EcoRI site and Escherichia coli transformants in which some translocation of beta-lactamase across the cytoplasmic membrane was restored were selected by their ability to survive and form colonies on plates containing a low level of ampicillin. About 15-20% of all in-frame inserts restored some beta-lactamase translocation and the salient feature of these sequences was their marked hydrophobicity. These results are discussed in the light of a similar study in which sequences able to function as translocators of invertase in yeast were cloned and analysed (Kaiser et al., 1987).
Mol Microbiol 1989 Oct
PMID:Identification of amino acid sequences that can function as translocators of beta-lactamase in Escherichia coli. 269 95

The scr genes located on plasmid pUR400 and responsible for sucrose (Scr) metabolism of Escherichia coli K12 and other enteric bacteria have been cloned on a 9.3 kb DNA fragment. The different genes were mapped by transposon insertion mutagenesis, by restriction endonuclease and deletion mapping, and the corresponding gene products were identified. Besides the known structural genes scrA, coding for an EnzymeII(Scr) (45 kD) of the phosphoenolypyruvate-dependent phosphotransferase system (PTS), and scrB, coding for a sucrose 6-phosphate hydrolase (invertase) (55 kD), two new structural genes were discovered. Gene scrK apparently codes for an intracellular and ATP-dependent fructokinase (39 kD), while scrY seems to code for a sucrose porin (58 kD) in the outer cell membrane. No genes for an Enzyme III(Scr) of the PTS or for (a) glycosyltransferase(s) were detected. The four genes form an scr operon (gene order, scrK scrY scrA scrB, transcription from K to B), regulated by a repressor (gene scrR, 37 kD) and inducible by sucrose, fructose and fructose-containing oligosaccharides.
Mol Microbiol 1988 Jan
PMID:Plasmid-mediated sucrose metabolism in Escherichia coli K12: mapping of the scr genes of pUR400. 283 84

In the yeast Saccharomyces cerevisiae six unlinked structural genes for invertase, the SUC genes, are known. We sequenced about 800 bp of the 5' non-coding region and the first 220 bp of the coding region of the genes SUC1, SUC3, SUC4 and SUC5 and compared them with the previously sequenced genes SUC2 and SUC7 (Sarokin and Carlson 1985a). All are highly homologous within the coding region but in the non-coding region SUC1 shows some differences and SUC2 is more highly diverged. Two different kinds of TATA boxes were identified: the more strongly expressed genes SUC1, 2 and 4 have the sequence TATAAA and the more weakly expressed genes SUC3, 5 and 7 have TACAAA. Though the SUC1 sequence is in general more homologous to the other SUC genes, the region between -140 and +100 of SUC1 is nearly identical to SUC2. This could be due to a gene conversion between SUC1 and the silent suc2 degrees allele which occurs in the strains carrying SUC1. Within the upstream regions of all the SUC genes three regions with palindromic sequences analogous to stem and loop structures were identified. Comparable structures could be detected in similar positions in the upstream sequences of the divergently transcribed yeast gene pairs MAL6S-MAL6T and GAL1-GAL10. Implications for the importance of these structures in the regulation and initiation of transcription are discussed.
Mol Gen Genet 1988 Mar
PMID:Structural analysis of the 5' regions of yeast SUC genes revealed analogous palindromes in SUC, MAL and GAL. 283 32

The SUC gene family of Saccharomyces contains six structural genes for invertase (SUC1 through SUC5 and SUC7) which are located on different chromosomes. Most yeast strains do not carry all six SUC genes and instead carry natural negative (suc0) alleles at some or all SUC loci. We determined the physical structures of SUC and suc0 loci. Except for SUC2, which is an unusual member of the family, all of the SUC genes are located very close to telomeres and are flanked by homologous sequences. On the centromere-proximal side of the gene, the conserved region contains X sequences, which are sequences found adjacent to telomeres (C. S. M. Chan and B.-K. Tye, Cell 33:563-573, 1983). On the other side of the gene, the homology includes about 4 kilobases of flanking sequence and then extends into a Y' element, which is an element often found distal to the X sequence at telomeres (Chan and Tye, Cell 33:563-573, 1983). Thus, these SUC genes and flanking sequences are embedded in telomere-adjacent sequences. Chromosomes carrying suc0 alleles (except suc20) lack SUC structural genes and portions of the conserved flanking sequences. The results indicate that the dispersal of SUC genes to different chromosomes occurred by rearrangements of chromosome telomeres.
Mol Cell Biol 1985 Nov
PMID:Evolution of the dispersed SUC gene family of Saccharomyces by rearrangements of chromosome telomeres. 301 85

Synthetic oligonucleotides coding for the yeast invertase secretion signal peptide were fused to the gene for the mature form of human interferon (huIFN-alpha 2). Two plasmids (E3 and F2) were constructed. E3 contained the invertase signal codons in a reading frame with the mature huIFN-alpha 2 gene. F2 had a deletion of the codon for alanine at amino acid residue-5 in the invertase signal and an addition of a methionine codon located between the coding sequences for the invertase signal and mature huIFN-alpha 2. Both hybrid genes were located adjacent to the promoter from the 3-phosphoglycerate kinase gene on the multicopy yeast expression plasmid, YEp1PT. Yeast transformants containing these plasmids produced somewhat more IFN than did the same expression plasmid containing the IFN gene with its human secretion signal sequence. HuIFN-alpha 2, purified from the medium of yeast cells containing E3, was found to be processed at the correct site. The huIFN-alpha 2 made by plasmid F2 was found to be completely processed at the junction between the invertase signal (a variant) and the methionine of methionine-huIFN-alpha 2. These results strongly suggested that the invertase signal (or its variant) attached to huIFN was efficiently recognized by the presumed signal recognition particle and was cleaved by the signal peptidase in the yeast cells. These results also suggested that amino acid changes on the right side of the cleavage site did not necessarily prevent cleavage or secretion.
Mol Cell Biol 1986 May
PMID:Saccharomyces cerevisiae secretes and correctly processes human interferon hybrid proteins containing yeast invertase signal peptides. 302 6

Using a selection for spontaneous mutants that mislocalize a vacuolar carboxypeptidase Y (CPY)-invertase fusion protein to the cell surface, we identified vacuolar protein targeting (vpt) mutants in 25 new vpt complementation groups. Additional alleles in each of the eight previously identified vpt complementation groups (vpt1 through vpt8) were also obtained. Representative alleles from each of the 33 vpt complementation groups (vpt1 through vpt33) were shown to exhibit defects in the sorting and processing of several native vacuolar proteins, including the soluble hydrolases CPY, proteinase A, and proteinase B. Of the 33 complementation groups, 19 were found to contain mutant alleles that led to extreme defects. In these mutants, CPY accumulated in its Golgi complex-modified precursor form which was secreted by the mutant cells. Normal protein secretion appeared to be unaffected in the vpt mutants. The lack of significant leakage of cytosolic markers from the vpt mutant cells indicated that the vacuolar protein-sorting defects associated with these mutants do not result from cell lysis. In addition, the observation that the precursor rather than the mature forms of CPY, proteinase A, proteinase B were secreted from the vpt mutants was consistent with the fact that mislocalization occurred at a stage after Golgi complex-specific modification, but before final vacuolar sorting of these enzymes. Vacuolar membrane protein sorting appeared to be unaffected in the majority of the vpt mutants. However, a subset of the vpt mutants (vpt11, vpt16, vpt18, and vpt33) was found to exhibit defects in the sorting of a vacuolar membrane marker enzyme, alpha-mannosidase. Up to 50% of the alpha-mannosidase enzyme activity was found to be mislocalized to the cell surface in these vpt mutants. Seven of the vpt complementation groups (vpt3, vpt11, vpt15, vpt16, vpt18, vpt29, and vpt33) contained alleles that led to a conditional lethal phenotype; the mutants were temperature sensitive for vegetative cell growth. This temperature-sensitive phenotype has been shown to be recessive and to cosegregate with the vacuolar protein-sorting defect in each case. Tetrad analysis showed that vpt3 mapped to the right arm of chromosome XV and that vpt15 mapped to the right arm of chromosome II. Intercrosses with other mutants that exhibited defects in vacuolar protein sorting or function (vpl, sec, pep, and end mutants) revealed several overlaps among these different sets of genes. Together, these data indicate that more than 50 gene products are involved, directly or indirectly, in the process of vacuolar protein sorting.
Mol Cell Biol 1988 Nov
PMID:Protein sorting in Saccharomyces cerevisiae: isolation of mutants defective in the delivery and processing of multiple vacuolar hydrolases. 306 74

Expression of secreted invertase from the SUC2 gene is regulated by carbon catabolite repression. Previously, an upstream regulatory region that is required for derepression of secreted invertase was identified and shown to confer glucose-repressible expression to the heterologous promoter of a LEU2-lacZ fusion. In this paper we show that tandem copies of a 32-base pair (bp) sequence from the upstream regulatory region activate expression of the same LEU2-lacZ fusion. The level of expression increased with the number of copies of the element, but was independent of their orientation; the expression from constructions containing four copies of the sequence was only twofold lower than that when the entire SUC2 upstream regulatory region was present. This activation was not significantly glucose repressible. The 32-bp sequence includes a 7-bp motif with the consensus sequence (A/C)(A/G)GAAAT that is repeated at five sites within the upstream regulatory region. Genetic evidence supporting the functional significance of this repeated motif was obtained by pseudoreversion of a SUC2 deletion mutant lacking part of the upstream region, including two copies of the 7-bp element. In three of five pseudorevertants, the mutations that restored high-level SUC2 expression altered one of the remaining copies of the 7-bp element.
Mol Cell Biol 1986 Jul
PMID:Short repeated elements in the upstream regulatory region of the SUC2 gene of Saccharomyces cerevisiae. 309 12

The structural gene for the enzyme levanase of Bacillus subtilis (SacC) was cloned in Escherichia coli. The cloned gene was mapped by PBS1 transduction near the sacL locus on the B. subtilis chromosome, between leuA and aroD. Expression of the enzyme was demonstrated both in B. subtilis and in E. coli. The presence of sacC allowed E. coli to grow on sucrose as the sole carbon source. The complete nucleotide sequence of sacC was determined. It includes an open reading frame of 2,031 bp, coding for a protein with calculated molecular weight of 75,866 Da, including a putative signal peptide similar to precursors of secreted proteins found in Bacilli. The apparent molecular weight of purified levanase is 73 kDa. The sacC gene product was characterized in an in vitro system and in a minicell-producing strain of E. coli, confirming the existence of a precursor form of levanase of about 75 kDa. Comparison of the predicted aminoacid sequence of levanase with those of the two other known beta-D-fructofuranosidases of B. subtilis indicated a homology with sucrase, but not with levansucrase. A stronger homology was detected with the N-terminal region of yeast invertase, suggesting the existence of a common ancestor.
Mol Gen Genet 1987 Jun
PMID:Characterization of the levanase gene of Bacillus subtilis which shows homology to yeast invertase. 311 19


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