scholarly journals Glu-370 in the Large Subunit Influences the Substrate Binding, Allosteric, and Heat Stability Properties of Potato ADP-glucose Pyrophosphorylase

2016 ◽  
Author(s):  
Ayse B Seferoglu ◽  
Seref Gul ◽  
Ugur M Dikbas ◽  
Kaan Koper ◽  
Ibrahim Baris ◽  
...  
Keyword(s):  
Computational Analysis ◽  
Large Subunit ◽  
Heat Stability ◽  
Starch Biosynthesis ◽  
Enzyme Kinetic ◽  
Wild Type ◽  
Binding Properties ◽  
Allosteric Enzyme ◽  
Heat Stable ◽  
Adp Glucose Pyrophosphorylase

ADP-glucose pyrophosphorylase (AGPase) is a key allosteric enzyme in plant starch biosynthesis. Plant AGPase is a heterotetrameric enzyme that consists of large (LS) and small subunits (SS), which are encoded by two different genes. In this study, we showed that the conversion of Glu to Gly at position 370 in the LS of AGPase alters the heterotetrameric stability along with the binding properties of substrate and effectors of the enzyme. Kinetic analyses revealed that the affinity of the LSE370GSSWT AGPase for glucose-1-phosphate is 3-fold less than for wild type (WT) AGPase. Additionally, the LSE370GSSWT AGPase requires 3-fold more 3-phosphogyceric acid to be activated. Finally, the LSE370GSSWTAGPase is less heat stable compared with the WT AGPase. Computational analysis of the mutant Gly-370 in the 3D modeled LS AGPase showed that this residue changes charge distribution of the surface and thus affect stability of the LS AGPase and overall heat stability of the heterotetrameric AGPase. In summary, our results show that LSE370 intricately modulate the heat stability and enzymatic activity of the AGPase.

A mutant of rice lacking the leaf large subunit of ADP-glucose pyrophosphorylase has drastically reduced leaf starch content but grows normally

2007 ◽  
Vol 34 (6) ◽  
pp. 480 ◽  
Author(s):  
Sandrine Rösti ◽  
Brendan Fahy ◽  
Kay Denyer
Keyword(s):  
Environmental Conditions ◽  
No Reduction ◽  
Starch Content ◽  
Large Subunit ◽  
Starch Synthesis ◽  
Small Subunit ◽  
Wild Type ◽  
Gene Encoding ◽  
Subunit Protein ◽  
Adp Glucose Pyrophosphorylase

A mutant of rice was identified with a Tos17 insertion in OsAPL1, a gene encoding a large subunit (LSU) of ADP-glucose pyrophosphorylase (AGPase). The insertion prevents production of a normal transcript from OsAPL1. Characterisation of the mutant (apl1) showed that the LSU encoded by OsAPL1 is required for AGPase activity in rice leaf blades. In mutant leaf blades, the AGPase small subunit protein is not detectable and the AGPase activity and starch content are reduced to <1 and <5% of that in wild type blades, respectively. The mutation also leads to a reduction in starch content in the leaf sheaths but does not significantly affect AGPase activity or starch synthesis in other parts of the plant. The sucrose, glucose and fructose contents of the leaves are not affected by the mutation. Despite the near absence of starch in the leaf blades, apl1 mutant rice plants grow and develop normally under controlled environmental conditions and show no reduction in productivity.

A second form of the single-strand specific endonuclease of Neurospora crassa which is associated with a double-strand exonuclease

1976 ◽  
Vol 54 (11) ◽  
pp. 971-980 ◽  
Author(s):  
M. J. Fraser ◽  
R. Tjeerde ◽  
K. Matsumoto
Keyword(s):  
Neurospora Crassa ◽  
Heat Stability ◽  
Single Strand ◽  
Dna Unwinding ◽  
Endonuclease Activity ◽  
Wild Type ◽  
Double Stranded Rna ◽  
Heat Stable ◽  
Double Stranded Dna ◽  
Heating Up

A second form of single-strand specific endonuclease, which is stable to heating up to 74 °C and does not bind strongly to phosphocellulose, has been partially purified from extracts of mycelia of wild-type Neurospora crassa. The endonuclease is associated with an equally heat-stable exonuclease which degrades linear but not circular double-stranded DNA and does not attack double-stranded RNA. The exonuclease probably also degrades single-stranded DNA. Both endonuclease and exonuclease activities are inhibited by 0.1–0.5 mM ATP. The exonuclease is preferentially inhibited by a variety of agents and preferentially inactivated by trypsin. A DNA-unwinding activity has also been detected in the nuclease preparation. Protease(s) present in the nuclease preparation destroy the DNA-unwinding and exonuclease activities on incubation at 37 °C, but do not affect the endonuclease activity. However, the heat-stability and chromatographic properties of the endonuclease are affected by this treatment. The altered properties of the endonuclease are very similar to those of the single-strand specific endonuclease which has been previously described. The combined nuclease activities of the unaltered preparation make up a putative recombination nuclease of N. crassa.

scholarly journals MULTIPLE FORMS OF MAIZE ENDOSPERM ADP-GLUCOSE PYROPHOSPHORYLASE AND THEIR CONTROL BY SHRUNKEN-2 AND BRITTLE-2

Genetics ◽  
1980 ◽  
Vol 95 (4) ◽  
pp. 961-970
Author(s):  
L C Hannah ◽  
D M Tuschall ◽  
R J Mans
Keyword(s):  
Total Activity ◽  
Stable Form ◽  
Wild Type ◽  
Multiple Forms ◽  
Maize Endosperm ◽  
Heat Stable ◽  
Heat Labile ◽  
Adp Glucose Pyrophosphorylase ◽  
Stable Forms

ABSTRACT Heat-labile and heat stable forms of ADP-glucose pyrophosphorylase were identified in the maize endosperm. The heat-labile form is destroyed by normal electrophoretic conditions. The heat-stable form corresponds to pyrophosphorylase B. In wild type, 96% of the total activity is heat labile. Both forms are reduced in 11 brittle-2 (bt2) and 12 shrunken-2 (sh2) mutants. The heat-labile form is reduced to a greater extent than is the heat-stable form in each of the 23 mutants. Deletion of sh2 abolishes both forms. The original ratio of the two forms is restored after sh2 function is expressed via transposition of Dissociation from sh2. The possible roles of these genes in the control of ADP-glucose pyrophosphorylase are discussed.

Intercellular transfer of beta-glucuronidase in chimeric mice

1979 ◽  
Vol 40 (1) ◽  
pp. 21-31
Author(s):  
K. Herrup ◽  
R.J. Mullen
Keyword(s):  
Gene Product ◽  
Heat Stability ◽  
Wild Type Allele ◽  
Chimeric Mice ◽  
Wild Type ◽  
Glucuronidase Activity ◽  
Heat Stable ◽  
Dark Staining ◽  
Activity Loss ◽  
Light Staining

Chimeric mice were produced by aggregating 2 embryos, each of which was homozygous for a different structural allele of the enzyme beta-glucuronidase. The alleles used were Gusb, the ‘wild-type’ allele, and Gush, an allele whose gene product shows decreased activity in all tissues as well as decreased heat stability. Staining of untreated adult chimeric livers for glucuronidase activity revealed a mosaic of high (Gusb) and low (Gush) activity cells. The boundaries between cells of different activity were sharp and revealed no diffusion of enzyme or reaction product. Treating sections at 73 degrees c before staining led to a decay of staining activity in normal (non-chimeric) Gush/Gush and Gusb/Gusb tissue. The rates of activity loss under the conditions used differed by 10-fold. Of the 2 genotypes in the chimera, the dark-staining, Gusb, cells decayed in a fashion similar to that of the Gusb/Gusb control. The light-staining, Gush, cells of the chimera lost their staining a a unique fashion. Within 20 min they quickly lost a majority of their staining activity but that which remained was relatively heat stable. The second Gusb-like phase of the decay, seen both photographically and photometrically, suggests that Gusb gene product has been transferred to cells of Gush/Gush genotype.

scholarly journals Glu-370 in the large subunit influences the substrate binding, allosteric, and heat stability properties of potato ADP-glucose pyrophosphorylase

Plant Science ◽  
2016 ◽  
Vol 252 ◽  
pp. 125-132
Author(s):  
Ayse Bengisu Seferoglu ◽  
Seref Gul ◽  
Ugur Meric Dikbas ◽  
Ibrahim Baris ◽  
Kaan Koper ◽  
...  
Keyword(s):  
Substrate Binding ◽  
Large Subunit ◽  
Heat Stability ◽  
Stability Properties ◽  
Adp Glucose Pyrophosphorylase

scholarly journals Transcriptional Activation in Yeast Cells Lacking Transcription Factor IIA

Genetics ◽  
1999 ◽  
Vol 153 (4) ◽  
pp. 1573-1581 ◽  
Author(s):  
Susanna Chou ◽  
Sukalyan Chatterjee ◽  
Mark Lee ◽  
Kevin Struhl
Keyword(s):  
Transcription Factor ◽  
Transcriptional Activation ◽  
Large Subunit ◽  
Yeast Cells ◽  
Specific Cell ◽  
Wild Type ◽  
Activation Domains ◽  
Cycle Arrest ◽  
General Transcription Factor

Abstract The general transcription factor IIA (TFIIA) forms a complex with TFIID at the TATA promoter element, and it inhibits the function of several negative regulators of the TATA-binding protein (TBP) subunit of TFIID. Biochemical experiments suggest that TFIIA is important in the response to transcriptional activators because activation domains can interact with TFIIA, increase recruitment of TFIID and TFIIA to the promoter, and promote isomerization of the TFIID-TFIIA-TATA complex. Here, we describe a double-shut-off approach to deplete yeast cells of Toa1, the large subunit of TFIIA, to &lt;1% of the wild-type level. Interestingly, such TFIIA-depleted cells are essentially unaffected for activation by heat shock factor, Ace1, and Gal4-VP16. However, depletion of TFIIA causes a general two- to threefold decrease of transcription from most yeast promoters and a specific cell-cycle arrest at the G2-M boundary. These results indicate that transcriptional activation in vivo can occur in the absence of TFIIA.

scholarly journals A procedure to introduce point mutations into the Rubisco large subunit gene in wild‐type plants

2021 ◽  
Author(s):  
Myat T. Lin ◽  
Douglas J. Orr ◽  
Dawn Worrall ◽  
Martin A. J. Parry ◽  
Elizabete Carmo‐Silva ◽  
...  
Keyword(s):  
Large Subunit ◽  
Point Mutations ◽  
Subunit Gene ◽  
Wild Type

scholarly journals A GENETICALLY DISTINCT FORM OF CYCLIC AMP PHOSPHODIESTERASE ASSOCIATED WITH CHROMOMERE 3D4 IN DROSOPHILA MELANOGASTER

Genetics ◽  
1979 ◽  
Vol 91 (3) ◽  
pp. 521-535
Author(s):  
John A Kiger ◽  
Eric Golanty
Keyword(s):  
Drosophila Melanogaster ◽  
Cyclic Amp ◽  
Structural Gene ◽  
Regulatory Gene ◽  
Heat Stability ◽  
Normal Female ◽  
Dependent Manner ◽  
Cyclic Amp Phosphodiesterase ◽  
Heat Stable ◽  
Camp Levels

ABSTRACT Two cyclic AMP phosphodiesterase enzymes (E.C.3.1.4.17) are present in homogenates of adult Drosophila melanogaster. The two enzymes differ from one another in heat stability, affinity for Mg++, Ca++ activation and molecular weight. They do not differ markedly in their affinities for cyclic AMP, and both exhibit anomalous Michaelis-Menten kinetics. The more heatlabile enzyme is controlled in a dosage-dependent manner by chromomere 3D4 of the X chromosome and is absent in flies that are deficient for chromomere 3D4. Chromomere 3D4 is also necessary for the maintenance of normal cAMP levels, for male fertility, and for normal female fertility and oogenesis. The structural gene(s) for the more heat-stable enzyme is located outside of chromomeres 3C12-3D4. Whether 3D4 contains a structural gene, or a regulatory gene necessary for the presence of the labile enzyme, remains to be determined.

scholarly journals Human Rev1 relies on insert-2 to promote selective binding and accurate replication of stabilized G-quadruplex motifs

2021 ◽  
Author(s):  
Amit Ketkar ◽  
Lane Smith ◽  
Callie Johnson ◽  
Alyssa Richey ◽  
Makayla Berry ◽  
...  
Keyword(s):  
Amino Acids ◽  
Mutation Frequency ◽  
Control Sequence ◽  
Wild Type ◽  
Binding Properties ◽  
High Affinity ◽  
G4 Dna ◽  
G Quadruplex ◽  
Forward Mutagenesis

Abstract We previously reported that human Rev1 (hRev1) bound to a parallel-stranded G-quadruplex (G4) from the c-MYC promoter with high affinity. We have extended those results to include other G4 motifs, finding that hRev1 exhibited stronger affinity for parallel-stranded G4 than either anti-parallel or hybrid folds. Amino acids in the αE helix of insert-2 were identified as being important for G4 binding. Mutating E466 and Y470 to alanine selectively perturbed G4 binding affinity. The E466K mutant restored wild-type G4 binding properties. Using a forward mutagenesis assay, we discovered that loss of hRev1 increased G4 mutation frequency &gt;200-fold compared to the control sequence. Base substitutions and deletions occurred around and within the G4 motif. Pyridostatin (PDS) exacerbated this effect, as the mutation frequency increased &gt;700-fold over control and deletions upstream of the G4 site more than doubled. Mutagenic replication of G4 DNA (±PDS) was partially rescued by wild-type and E466K hRev1. The E466A or Y470A mutants failed to suppress the PDS-induced increase in G4 mutation frequency. These findings have implications for the role of insert-2, a motif conserved in vertebrates but not yeast or plants, in Rev1-mediated suppression of mutagenesis during G4 replication.

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