scholarly journals Structure, Function and Regulation of a Second Pyruvate Kinase Isozyme in Pseudomonas aeruginosa

2021 ◽  
Vol 12 ◽  
Author(s):  
Yassmin Abdelhamid ◽  
Meng Wang ◽  
Susannah L. Parkhill ◽  
Paul Brear ◽  
Xavier Chee ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Structure Function ◽  
Pyruvate Kinase ◽  
Amino Acid Sequence Identity ◽  
Conformational Transition ◽  
Allosteric Activation ◽  
X Ray ◽  
Directed Mutation ◽  
Sequence Identity ◽  
Allosteric Regulator

Pseudomonas aeruginosa (PA) depends on the Entner-Doudoroff pathway (EDP) for glycolysis. The main enzymatic regulator in the lower half of the EDP is pyruvate kinase. PA contains genes that encode two isoforms of pyruvate kinase, denoted PykAPA and PykFPA. In other well-characterized organisms containing two pyruvate kinase isoforms (such as Escherichia coli) each isozyme is differentially regulated. The structure, function and regulation of PykAPA has been previously characterized in detail, so in this work, we set out to assess the biochemical and structural properties of the PykFPA isozyme. We show that pykFPA expression is induced in the presence of the diureide, allantoin. In spite of their relatively low amino acid sequence identity, PykAPA and PykFPA display broadly comparable kinetic parameters, and are allosterically regulated by a very similar set of metabolites. However, the x-ray crystal structure of PykFPA revealed significant differences compared with PykAPA. Notably, although the main allosteric regulator binding-site of PykFPA was empty, the “ring loop” covering the site adopted a partially closed conformation. Site-directed mutation of the proline residues flanking the ring loop yielded apparent “locked on” and “locked off” allosteric activation phenotypes, depending on the residue mutated. Analysis of PykFPA inter-protomer interactions supports a model in which the conformational transition(s) accompanying allosteric activation involve re-orientation of the A and B domains of the enzyme and subsequent closure of the active site.

scholarly journals Evolutionary plasticity in the allosteric regulator-binding site of pyruvate kinase isoform PykA from Pseudomonas aeruginosa

2019 ◽  
Vol 294 (42) ◽  
pp. 15505-15516 ◽  
Author(s):  
Yassmin Abdelhamid ◽  
Paul Brear ◽  
Jack Greenhalgh ◽  
Xavier Chee ◽  
Taufiq Rahman ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Binding Site ◽  
Pyruvate Kinase ◽  
Allosteric Regulator

scholarly journals Changes in the allosteric site of human liver pyruvate kinase upon activator binding include the breakage of an intersubunit cation–π bond

2019 ◽  
Vol 75 (6) ◽  
pp. 461-469 ◽  
Author(s):  
Jeffrey S. McFarlane ◽  
Trey A. Ronnebaum ◽  
Kathleen M. Meneely ◽  
Annemarie Chilton ◽  
Aron W. Fenton ◽  
...  
Keyword(s):  
Pyruvate Kinase ◽  
Functional Data ◽  
Human Liver ◽  
Phosphate Binding ◽  
Allosteric Site ◽  
Allosteric Activation ◽  
X Ray ◽  
Fructose 1,6 Bisphosphate ◽  
Open Position ◽  
Binding Loop

Human liver pyruvate kinase (hLPYK) converts phosphoenolpyruvate to pyruvate in the final step of glycolysis. hLPYK is allosterically activated by fructose-1,6-bisphosphate (Fru-1,6-BP). The allosteric site, as defined by previous structural studies, is located in domain C between the phosphate-binding loop (residues 444–449) and the allosteric loop (residues 527–533). In this study, the X-ray crystal structures of four hLPYK variants were solved to make structural correlations with existing functional data. The variants are D499N, W527H, Δ529/S531G (called GGG here) and S531E. The results revealed a conformational toggle between the open and closed positions of the allosteric loop. In the absence of Fru-1,6-BP the open position is stabilized, in part, by a cation–π bond between Trp527 and Arg538′ (from an adjacent monomer). In the S531E variant glutamate binds in place of the 6′-phosphate of Fru-1,6-BP in the allosteric site, leading to partial allosteric activation. Finally, the structure of the D499N mutant does not provide structural evidence for the previously observed allosteric activation of the D499N variant.

Heterogeneity in hydrophobic deep eutectic solvents: SAXS prepeak and local environments

2021 ◽  
Vol 23 (6) ◽  
pp. 3915-3924
Author(s):  
Akshay Malik ◽  
Hemant K. Kashyap
Keyword(s):  
Structure Function ◽  
Deep Eutectic Solvents ◽  
Structural Heterogeneity ◽  
X Ray ◽  
Intermediate Range ◽  
X Ray Scattering ◽  
Local Environments ◽  
Ray Scattering

The observation of the prepeak in the simulated total X-ray scattering structure function (S(q)) reveals the presence of intermediate-range structural heterogeneity in hydrophobic deep eutectic solvents.

scholarly journals Exploring the “minimal” structure of a functional ADAMTS13 by mutagenesis and small-angle X-ray scattering

Blood ◽  
2019 ◽  
Vol 133 (17) ◽  
pp. 1909-1918 ◽  
Author(s):  
Jian Zhu ◽  
Joshua Muia ◽  
Garima Gupta ◽  
Lisa A. Westfield ◽  
Karen Vanhoorelbeke ◽  
...  
Keyword(s):  
Small Angle ◽  
Columba Livia ◽  
Allosteric Activation ◽  
Complement Components ◽  
X Ray ◽  
X Ray Scattering ◽  
Morphogenetic Protein ◽  
Minimal Structure ◽  
Von Willebrand ◽  
Ray Scattering

Abstract Human ADAMTS13 is a multidomain protein with metalloprotease (M), disintegrin-like (D), thrombospondin-1 (T), Cys-rich (C), and spacer (S) domains, followed by 7 additional T domains and 2 CUB (complement components C1r and C1s, sea urchin protein Uegf, and bone morphogenetic protein-1) domains. ADAMTS13 inhibits the growth of von Willebrand factor (VWF)–platelet aggregates by cleaving the cryptic Tyr1605-Met1606 bond in the VWF A2 domain. ADAMTS13 is regulated by substrate-induced allosteric activation; without shear stress, the distal T8-CUB domains markedly inhibit VWF cleavage, and binding of VWF domain D4 or selected monoclonal antibodies (MAbs) to distal ADAMTS13 domains relieves this autoinhibition. By small angle X-ray scattering (SAXS), ADAMTS13 adopts a hairpin-like conformation with distal T7-CUB domains close to the proximal MDTCS domains and a hinge point between T4 and T5. The hairpin projects like a handle away from the core MDTCS and T7-CUB complex and contains distal T domains that are dispensable for allosteric regulation. Truncated constructs that lack the T8-CUB domains are not autoinhibited and cannot be activated by VWF D4 but retain the hairpin fold. Allosteric activation by VWF D4 requires T7, T8, and the 58–amino acid residue linker between T8 and CUB1. Deletion of T3 to T6 produced the smallest construct (delT3-6) examined that could be activated by MAbs and VWF D4. Columba livia (pigeon) ADAMTS13 (pADAMTS13) resembles human delT3-6, retains normal activation by VWF D4, and has a SAXS envelope consistent with amputation of the hairpin containing the dispensable T domains of human ADAMTS13. Our findings suggest that human delT3-6 and pADAMTS13 approach a “minimal” structure for allosterically regulated ADAMTS13.

Biochemical characterization of a glucoamylase from Saccharomycopsis fibuligera R64

Biologia ◽  
2011 ◽  
Vol 66 (1) ◽  
Author(s):  
Dessy Natalia ◽  
Keni Vidilaseris ◽  
Pasjan Satrimafitrah ◽  
Wangsa Ismaya ◽  
Purkan ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Amino Acid ◽  
Amino Acid Sequence Identity ◽  
Biochemical Characterization ◽  
Saccharomycopsis Fibuligera ◽  
Sequence Identity ◽  
Ic50 Value ◽  
High Amino Acid ◽  
Ph Range

AbstractGlucoamylase from the yeast Saccharomycopsis fibuligera R64 (GLL1) has successfully been purified and characterized. The molecular mass of the enzyme was 56,583 Da as determined by mass spectrometry. The purified enzyme demonstrated optimum activity in the pH range of 5.6–6.4 and at 50°C. The activity of the enzyme was inhibited by acarbose with the IC50 value of 5 μM. GLL1 shares high amino acid sequence identity with GLU1 and GLA1, which are Saccharomycopsis fibuligera glucoamylases from the strains HUT7212 and KZ, respectively. The properties of GLL1, however, resemble that of GLU1. The elucidation of the primary structure of GLL1 contributes to the explanation of this finding.

scholarly journals Petunia vein banding virus: Characterization of a New Tymovirus from Petunia × hybrida

Plant Disease ◽  
2000 ◽  
Vol 84 (7) ◽  
pp. 739-742 ◽  
Author(s):  
M. A. V. Alexandre ◽  
L. M. L. Duarte ◽  
E. B. Rivas ◽  
C. M. Chagas ◽  
M. M. Barradas ◽  
...  
Keyword(s):  
Nicotiana Benthamiana ◽  
Petunia Hybrida ◽  
Amino Acid Sequence Identity ◽  
Rio Grande Do Sul ◽  
Cytopathic Effects ◽  
Sequence Identity ◽  
Andean Potato ◽  
Plant Families ◽  
Nicandra Physalodes

Petunia plants from a nursery in the State of Rio Grande do Sul, Brazil, showed pronounced vein banding and contained isometric particles with diameters of approximately 45 and 30 nm. The larger ones apparently represent a caulimovirus, while the smaller ones, which included both empty shells and full particles, were identified as those of a new tymovirus for which we propose the name Petunia vein banding virus (PetVBV). Originally, PetVBV was transmitted only with difficulty to healthy petunia plants. However, from an experimentally infected petu-nia, it was later readily transmitted also to Nicotiana benthamiana and Nicandra physalodes, but not to other species in the Solanaceae or other plant families. It produces cytopathic effects typical for tymovirus infections. Its coat protein shows approximately 65% amino acid sequence identity with those of Eggplant mosaic and Andean potato latent viruses, to which it is also serologically more closely related than to any other tymoviruses.

scholarly journals X-ray Irradiated Vaccine Confers protection against Pneumonia caused by Pseudomonas Aeruginosa

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Yanyan Li ◽  
Zhenling Wang ◽  
Xiaoxiao Liu ◽  
Jianying Tang ◽  
Bin Peng ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
X Ray

X-Ray Crystallographic Studies of the Structure-Function Relationships of HIV-1 Protease

1998 ◽  
pp. 59-63
Author(s):  
Lin Hong ◽  
Cai Zhang ◽  
Jean A. Hartsuck ◽  
Steve Foundling ◽  
Jordan Tang
Keyword(s):  
Structure Function ◽  
X Ray ◽  
Hiv 1
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