scholarly journals The β-amylase7 gene in Zea mays encodes a protein with structural and catalytic properties similar to Arabidopsis BAM2

2021 ◽  
Author(s):  
Claire M Ravenburg ◽  
McKayla B Riney ◽  
Jonathan D Monroe ◽  
Christopher E Berndsen
Keyword(s):  
Solution Structure ◽  
Short Form ◽  
Green Plant ◽  
Scattering Data ◽  
Sequence Alignments ◽  
E Coli ◽  
X Ray Scattering ◽  
Catalytically Active ◽  
Sigmoidal Kinetics ◽  
Functionally Diverse

Starch accumulates in the plastids of green plant tissue during the day to provide carbon for metabolism at night. Starch hydrolysis is catalyzed by members of the β-amylase (BAM) family, which in Arabidopsis thaliana (At), includes nine structurally and functionally diverse members. One of these enzymes, AtBAM2, is a plastid-localized enzyme that is unique among characterized β-amylases since it is tetrameric and exhibits sigmoidal kinetics. Sequence alignments show that the BAM domains of AtBAM7, a catalytically inactive, nuclear-localized transcription factor with an N-terminal DNA binding domain, and AtBAM2 are more closely related to each other than they are to any other AtBAM. Since BAM2 is found in more ancient lineages, it was hypothesized that BAM7 evolved from BAM2. However, analysis of the genomes of 48 flowering plants revealed 12 species that appear to have a BAM7 gene but lack a BAM2 gene. Upon closer inspection, these BAM7 proteins have a greater percent identity to AtBAM2 than to AtBAM7, and they share all of the AtBAM2 functional residues that BAM7 proteins normally lack. We hypothesize that these genes may encode a BAM2-like protein although they are currently annotated as BAM7-like genes. To test this hypothesis, we designed a cDNA of the short form of corn BAM7 (ZmBAM7-S) for expression in E. coli. Small Angle X-Ray Scattering data indicate that ZmBAM7-S has a tetrameric solution structure more similar to that of AtBAM2 than AtBAM1. In addition, partially purified ZmBAM7-S is catalytically active and exhibits sigmoidal kinetics. Together these data suggest that some BAM7 genes may encode a functional BAM2. Exploring and understanding β-amylase gene structure could have impacts on the current annotation of genes.

scholarly journals Low-resolution structure of the soluble domain GPAA1 (yGPAA170–247) of the glycosylphosphatidylinositol transamidase subunit GPAA1 from Saccharomyces cerevisiae

2013 ◽  
Vol 33 (2) ◽  
Author(s):  
Wuan Geok Saw ◽  
Birgit Eisenhaber ◽  
Frank Eisenhaber ◽  
Gerhard Grüber
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Solution Structure ◽  
Scattering Data ◽  
Radius Of Gyration ◽  
Low Resolution ◽  
X Ray Scattering ◽  
Eukaryotic Proteins ◽  
Structural Content ◽  
Α Helix ◽  
Gpi Transamidase

The GPI (glycosylphosphatidylinositol) transamidase complex catalyses the attachment of GPI anchors to eukaryotic proteins in the lumen of ER (endoplasmic reticulum). The Saccharomyces cerevisiae GPI transamidase complex consists of the subunits yPIG-K (Gpi8p), yPIG-S (Gpi17p), yPIG-T (Gpi16p), yPIG-U (CDC91/GAB1) and yGPAA1. We present the production of the two recombinant proteins yGPAA170–247 and yGPAA170–339 of the luminal domain of S. cerevisiae GPAA1, covering the amino acids 70–247 and 70–339 respectively. The secondary structural content of the stable and monodisperse yGPAA170–247 has been determined to be 28% α-helix and 27% β-sheet. SAXS (small-angle X-ray scattering) data showed that yGPAA170–247 has an Rg (radius of gyration) of 2.72±0.025 nm and Dmax (maximum dimension) of 9.14 nm. These data enabled the determination of the two domain low-resolution solution structure of yGPAA170–247. The large elliptical shape of yGPAA170–247 is connected via a short stalk to the smaller hook-like domain of 0.8 nm in length and 3.5 nm in width. The topological arrangement of yGPAA170–247 will be discussed together with the recently determined low-resolution structures of yPIG-K24–337 and yPIG-S38–467 from S. cerevisiae in the GPI transamidase complex.

scholarly journals Addressing the characterisation challenge to understand catalysis in MOFs: the case of nanoscale Cu supported in NU-1000

2017 ◽  
Vol 201 ◽  
pp. 337-350 ◽  
Author(s):  
Ana E. Platero-Prats ◽  
Zhanyong Li ◽  
Leighanne C. Gallington ◽  
Aaron W. Peters ◽  
Joseph T. Hupp ◽  
...  
Keyword(s):  
Dynamic Structure ◽  
Atomic Layer ◽  
Scattering Data ◽  
Catalyst Activation ◽  
Reducing Atmosphere ◽  
X Ray Scattering ◽  
Layer Deposition ◽  
Pdf Analysis ◽  
Catalytically Active

We explore the dynamic structure and reactivity of Cu species supported on NU-1000. By combining pair distribution function (PDF) analysis and difference envelope density (DED) analysis of in situ synchrotron-based X-ray scattering data, we simultaneously probe the local structure of supported Cu-species, their distribution within NU-1000 and distortions of the NU-1000 lattice under conditions relevant to catalysis and catalyst activation. These analyses show that atomic layer deposition (ALD) of Cu in NU-1000 (Cu-AIM) leads to the formation of Cu-oxo clusters within the small pores that connect the triangular and hexagonal channels. Exposure of Cu-AIM to a reducing atmosphere at 200 °C produces metallic Cu0 of two distinct particle sizes: ∼4 nm nanoparticles and small sub-nanometer clusters. The size of these nanoparticles appears to be constrained by NU-1000 pore dimensions, with evidence of the sub-nanometer clusters being bound within the triangular channels flanked by pyrene rings. This supported Cu0–NU-1000 system is catalytically active for gas-phase ethylene hydrogenation. Exposure of the catalyst to oxidative atmosphere re-oxidises the Cu species to a Cu2O cuprite phase. The dynamic restructuring of the system in different chemical environments underscores the importance of probing these systems in situ.

scholarly journals Solution structure and assembly of β-amylase2 from Arabidopsis thaliana

2019 ◽  
Author(s):  
Nithesh P. Chandrasekharan ◽  
Claire M. Ravenburg ◽  
Ian R. Roy ◽  
Jonathan D. Monroe ◽  
Christopher E. Berndsen
Keyword(s):  
Arabidopsis Thaliana ◽  
Ipomoea Batatas ◽  
Solution Structure ◽  
Scattering Data ◽  
Controlled Synthesis ◽  
X Ray ◽  
Domain Structures ◽  
X Ray Scattering ◽  
Loop Sequence ◽  
Assembly Analysis

AbstractStarch is a key energy storage molecule in plants that requires controlled synthesis and breakdown for effective plant growth. β-amylases (BAMs) hydrolyze starch into maltose to help meet the metabolic needs of the plant. In the model plant, Arabidopsis thaliana, there are nine BAMs which have apparently distinct functional and domain structures, although the functions of only a few of the BAMs are known and there are no 3-D structures of BAMs from this organism. Recently, AtBAM2 was proposed to form a tetramer based on chromatography and activity assays of mutants, however there was no direct observation of this tetramer. We collected small-angle X-ray scattering data on AtBAM2 and N-terminal mutants to describe the structure and assembly of the tetramer. Comparison of the scattering of the AtBAM2 tetramer to data collected using the sweet potato (Ipomoea batatas) BAM5, which is also reported to form a tetramer, showed there were differences in the overall assembly. Analysis of N-terminal truncations of AtBAM2 identified a loop sequence found only in BAM2 orthologs that appears to be critical for AtBAM2 tetramer assembly as well as activity.

scholarly journals Lactoferricins access the cytosol of Escherichia coli within few seconds

2021 ◽  
Author(s):  
Enrico Federico Semeraro ◽  
Lisa Marx ◽  
Johannes Mandl ◽  
Ilse Letofsky-Papst ◽  
Claudia Mayrhofer ◽  
...  
Keyword(s):  
Cell Envelope ◽  
Scattering Data ◽  
Bacterial Killing ◽  
Time Resolved ◽  
E Coli ◽  
Multi Scale ◽  
X Ray Scattering ◽  
Membrane Barrier ◽  
Collateral Effect ◽  
Peptide Partitioning

We report the real-time response of E. coli to lactoferricin-derived antimicrobial peptides (AMPs) on length-scales bridging microscopic cell-sizes to nanoscopic lipid packing using millisecond time-resolved synchrotron small-angle X-ray scattering. Coupling a multi-scale scattering data analysis to biophysical assays for peptide partitioning revealed that the AMPs rapidly saturate the bacterial envelope and reach the cytosol within less than three seconds—much faster than previously considered. Final cytosolic AMP concentrations of ~ 100 mM suggest an efficient shut-down of metabolism as primary cause for bacterial killing. On the other hand, the damage of the cell envelope is a collateral effect of AMP activity that does not kill the bacteria. This implies that the impairment of the membrane barrier is a necessary but not sufficient condition for microbial killing by lactoferricins. The most efficient AMP studied exceeds others in both speed of reaching cytoplasm and lowest cytosolic peptide concentration.

scholarly journals The solution structure of the complement deregulator FHR5 reveals a compact dimer and provides new insights into CFHR5 nephropathy

2020 ◽  
Vol 295 (48) ◽  
pp. 16342-16358
Author(s):  
Nilufar Kadkhodayi-Kholghi ◽  
Jayesh S. Bhatt ◽  
Jayesh Gor ◽  
Lindsay C. McDermott ◽  
Daniel P. Gale ◽  
...  
Keyword(s):  
Solution Structure ◽  
Factor H ◽  
Scattering Data ◽  
Radius Of Gyration ◽  
Domain Pair ◽  
Complement Factor ◽  
X Ray ◽  
X Ray Scattering ◽  
Complement Regulator ◽  
Ray Scattering

The human complement Factor H–related 5 protein (FHR5) antagonizes the main circulating complement regulator Factor H, resulting in the deregulation of complement activation. FHR5 normally contains nine short complement regulator (SCR) domains, but a FHR5 mutant has been identified with a duplicated N-terminal SCR-1/2 domain pair that causes CFHR5 nephropathy. To understand how this duplication causes disease, we characterized the solution structure of native FHR5 by analytical ultracentrifugation and small-angle X-ray scattering. Sedimentation velocity and X-ray scattering indicated that FHR5 was dimeric, with a radius of gyration (Rg) of 5.5 ± 0.2 nm and a maximum protein length of 20 nm for its 18 domains. This result indicated that FHR5 was even more compact than the main regulator Factor H, which showed an overall length of 26–29 nm for its 20 SCR domains. Atomistic modeling for FHR5 generated a library of 250,000 physically realistic trial arrangements of SCR domains for scattering curve fits. Only compact domain structures in this library fit well to the scattering data, and these structures readily accommodated the extra SCR-1/2 domain pair present in CFHR5 nephropathy. This model indicated that mutant FHR5 can form oligomers that possess additional binding sites for C3b in FHR5. We conclude that the deregulation of complement regulation by the FHR5 mutant can be rationalized by the enhanced binding of FHR5 oligomers to C3b deposited on host cell surfaces. Our FHR5 structures thus explained key features of the mechanism and pathology of CFHR5 nephropathy.

Small-Angle X-ray Scattering Reveals the Solution Structure of a Bacteriophytochrome in the Catalytically Active Pr State

2006 ◽  
Vol 364 (4) ◽  
pp. 655-666 ◽  
Author(s):  
Katie Evans ◽  
J. Günter Grossmann ◽  
Anthony P. Fordham-Skelton ◽  
Miroslav Z. Papiz
Keyword(s):  
Small Angle ◽  
Solution Structure ◽  
X Ray ◽  
X Ray Scattering ◽  
Catalytically Active ◽  
Ray Scattering

scholarly journals Solution structure and assembly of β-amylase 2 from Arabidopsis thaliana

2020 ◽  
Vol 76 (4) ◽  
pp. 357-365 ◽  
Author(s):  
Nithesh P. Chandrasekharan ◽  
Claire M. Ravenburg ◽  
Ian R. Roy ◽  
Jonathan D. Monroe ◽  
Christopher E. Berndsen
Keyword(s):  
Arabidopsis Thaliana ◽  
Ipomoea Batatas ◽  
Solution Structure ◽  
Scattering Data ◽  
Controlled Synthesis ◽  
X Ray ◽  
Domain Structures ◽  
X Ray Scattering ◽  
Loop Sequence ◽  
Assembly Analysis

Starch is a key energy-storage molecule in plants that requires controlled synthesis and breakdown for effective plant growth. β-Amylases (BAMs) hydrolyze starch into maltose to help to meet the metabolic needs of the plant. In the model plant Arabidopsis thaliana there are nine BAMs, which have apparently distinct functional and domain structures, although the functions of only a few of the BAMs are known and there are no 3D structures of BAMs from this organism. Recently, AtBAM2 was proposed to form a tetramer based on chromatography and activity assays of mutants; however, there was no direct observation of this tetramer. Here, small-angle X-ray scattering data were collected from AtBAM2 and its N-terminal truncations to describe the structure and assembly of the tetramer. Comparison of the scattering of the AtBAM2 tetramer with data collected from sweet potato (Ipomoea batatas) BAM5, which is also reported to form a tetramer, showed there were differences in the overall assembly. Analysis of the N-terminal truncations of AtBAM2 identified a loop sequence found only in BAM2 orthologs that appears to be critical for AtBAM2 tetramer assembly as well as for activity.

scholarly journals Structure of the amorphous titania precursor phase of N-doped photocatalysts

RSC Advances ◽  
2021 ◽  
Vol 11 (15) ◽  
pp. 8619-8627
Author(s):  
I. E. Grey ◽  
P. Bordet ◽  
N. C. Wilson
Keyword(s):  
Thermal Analyses ◽  
Real Space ◽  
Ammonia Solution ◽  
Scattering Data ◽  
X Ray ◽  
X Ray Scattering ◽  
Amorphous Titania ◽  
Precursor Phase ◽  
Titania Precursor ◽  
Titanyl Sulphate

Amorphous titania samples prepared by ammonia solution neutralization of titanyl sulphate have been characterized by chemical and thermal analyses, and with reciprocal-space and real-space fitting of wide-angle synchrotron X-ray scattering data.

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