scholarly journals The Molecular Basis of Sulfosugar Selectivity in Sulfoglycolysis

2021 ◽  
Author(s):  
Mahima Sharma ◽  
Palika Abayakoon ◽  
Ruwan Epa ◽  
Yi Jin ◽  
James P. Lingford ◽  
...  
Keyword(s):  
Biochemical Analysis ◽  
Dihydroxyacetone Phosphate ◽  
Central Metabolism ◽  
Photosynthetic Organisms ◽  
Catalytically Active ◽  
Glycolysis Pathway ◽  
Binding Pockets ◽  
Superficial Similarity ◽  
Complex Modulation ◽  
Mechanism Specificity

The sulfosugar sulfoquinovose (SQ) is produced by essentially all photosynthetic organisms on earth and is metabolized by bacteria through the process of sulfoglycolysis. The sulfoglycolytic Embden-Meyerhof-Parnas pathway metabolises SQ to produce dihydroxyacetone phosphate and sulfolactaldehyde and is analogous to the classical Embden-Meyerhof-Parnas glycolysis pathway for the metabolism of glucose-6-phosphate, though the former only provides one C3 fragment to central metabolism, with excretion of the other C3 fragment as dihydroxypropanesulfonate. Here, we report a comprehensive structural and biochemical analysis of the three core steps of sulfoglycolysis catalyzed by SQ isomerase, sulfofructose (SF) kinase and sulfofructose-1-phosphate (SFP) aldolase. Our data shows that despite the superficial similarity of this pathway to glycolysis, the sulfoglycolytic enzymes are specific for SQ metabolites and are not catalytically active on related metabolites from glycolytic pathways. This observation is rationalized by 3D structures of each enzyme, which reveal the presence of conserved sulfonate-binding pockets. We show that SQ isomerase acts preferentially on the β-anomer of SQ and reversibly produces both SF and sulforhamnose (SR), a previously unknown sugar that acts as a transcriptional regulator for the transcriptional repressor CsqR that regulates SQ-utilization. We also demonstrate that SF kinase is a key regulatory enzyme for the pathway that experiences complex modulation by the metabolites AMP, ADP, ATP, F6P, FBP, PEP, and citrate, and we show that SFP aldolase reversibly synthesizes SFP. This body of work provides fresh insights into the mechanism, specificity and regulation of sulfoglycolysis and has important implications for understanding how this biochemistry interfaces with central metabolism in prokaryotes to process this major repository of biogeochemical sulfur.

scholarly journals The Molecular Basis of Sulfosugar Selectivity in Sulfoglycolysis

2020 ◽  
Author(s):  
Mahima Sharma ◽  
Palika Abayakoon ◽  
Yi Jin ◽  
Ruwan Epa ◽  
James P. Lingford ◽  
...  
Keyword(s):  
Biochemical Analysis ◽  
Allosteric Modulation ◽  
Dihydroxyacetone Phosphate ◽  
Central Metabolism ◽  
Photosynthetic Organisms ◽  
Catalytically Active ◽  
Glycolysis Pathway ◽  
Binding Pockets ◽  
Superficial Similarity ◽  
Mechanism Specificity

The sulfosugar sulfoquinovose (SQ) is produced by essentially all photosynthetic organisms on earth and is metabolized bacteria through the process of sulfoglycolysis. The sulfoglycolytic Embden-Meyerhof-Parnas pathway metabolises SQ to produce dihydroxyacetone phosphate and sulfolactaldehyde and is analogous to the classical Embden-Meyerhof-Parnas glycolysis pathway for the metabolism of glucose-6-phosphate, though the former only provides one C3 fragment to central metabolism, with excretion of the other C3 fragment as dihydroxypropanesulfonate. Here, we report a comprehensive structural and biochemical analysis of the three core steps of sulfoglycolysis catalyzed by SQ isomerase, sulfofructose (SF) kinase and sulfofructose-1-phosphate aldolase. Our data shows that despite the superficial similarity of this pathway to glycolysis, the sulfoglycolytic enzymes are specific for SQ metabolites and are not catalytically active on related metabolites from glycolytic pathways. This observation is rationalized by 3D structures of each of these enzymes, which reveal the presence of conserved sulfonate-binding pockets. We show that SQ isomerase acts preferentially on the b-anomer of SQ and reversibly produces both SF and sulforhamnose (SR), a previously unknown sugar that acts as a transcriptional regulator for the transcriptional repressor CsqR that regulates SQ-utilisation. We also demonstrate that SF kinase is a key regulatory enzyme for the pathway that experiences complex allosteric modulation by the metabolites AMP, ADP, ATP, F6P, FBP, PEP, and citrate. This body of work provides fresh insights into the mechanism, specificity and regulation of sulfoglycolysis and has important implications for understanding how this biochemistry interfaces with central metabolism in prokaryotes to process this major repository of biogeochemical sulfur.

scholarly journals The Molecular Basis of Sulfosugar Selectivity in Sulfoglycolysis

2020 ◽  
Author(s):  
Mahima Sharma ◽  
Palika Abayakoon ◽  
Yi Jin ◽  
Ruwan Epa ◽  
James P. Lingford ◽  
...  
Keyword(s):  
Biochemical Analysis ◽  
Allosteric Modulation ◽  
Dihydroxyacetone Phosphate ◽  
Central Metabolism ◽  
Photosynthetic Organisms ◽  
Catalytically Active ◽  
Glycolysis Pathway ◽  
Binding Pockets ◽  
Superficial Similarity ◽  
Mechanism Specificity

The sulfosugar sulfoquinovose (SQ) is produced by essentially all photosynthetic organisms on earth and is metabolized bacteria through the process of sulfoglycolysis. The sulfoglycolytic Embden-Meyerhof-Parnas pathway metabolises SQ to produce dihydroxyacetone phosphate and sulfolactaldehyde and is analogous to the classical Embden-Meyerhof-Parnas glycolysis pathway for the metabolism of glucose-6-phosphate, though the former only provides one C3 fragment to central metabolism, with excretion of the other C3 fragment as dihydroxypropanesulfonate. Here, we report a comprehensive structural and biochemical analysis of the three core steps of sulfoglycolysis catalyzed by SQ isomerase, sulfofructose (SF) kinase and sulfofructose-1-phosphate aldolase. Our data shows that despite the superficial similarity of this pathway to glycolysis, the sulfoglycolytic enzymes are specific for SQ metabolites and are not catalytically active on related metabolites from glycolytic pathways. This observation is rationalized by 3D structures of each of these enzymes, which reveal the presence of conserved sulfonate-binding pockets. We show that SQ isomerase acts preferentially on the b-anomer of SQ and reversibly produces both SF and sulforhamnose (SR), a previously unknown sugar that acts as a transcriptional regulator for the transcriptional repressor CsqR that regulates SQ-utilisation. We also demonstrate that SF kinase is a key regulatory enzyme for the pathway that experiences complex allosteric modulation by the metabolites AMP, ADP, ATP, F6P, FBP, PEP, and citrate. This body of work provides fresh insights into the mechanism, specificity and regulation of sulfoglycolysis and has important implications for understanding how this biochemistry interfaces with central metabolism in prokaryotes to process this major repository of biogeochemical sulfur.

scholarly journals The Molecular Basis of Sulfosugar Selectivity in Sulfoglycolysis

2020 ◽  
Author(s):  
Mahima Sharma ◽  
Palika Abayakoon ◽  
Yi Jin ◽  
Ruwan Epa ◽  
James P. Lingford ◽  
...  
Keyword(s):  
Biochemical Analysis ◽  
Allosteric Modulation ◽  
Dihydroxyacetone Phosphate ◽  
Central Metabolism ◽  
Photosynthetic Organisms ◽  
Catalytically Active ◽  
Glycolysis Pathway ◽  
Binding Pockets ◽  
Superficial Similarity ◽  
Mechanism Specificity

The sulfosugar sulfoquinovose (SQ) is produced by essentially all photosynthetic organisms on earth and is metabolized bacteria through the process of sulfoglycolysis. The sulfoglycolytic Embden-Meyerhof-Parnas pathway metabolises SQ to produce dihydroxyacetone phosphate and sulfolactaldehyde and is analogous to the classical Embden-Meyerhof-Parnas glycolysis pathway for the metabolism of glucose-6-phosphate, though the former only provides one C3 fragment to central metabolism, with excretion of the other C3 fragment as dihydroxypropanesulfonate. Here, we report a comprehensive structural and biochemical analysis of the three core steps of sulfoglycolysis catalyzed by SQ isomerase, sulfofructose (SF) kinase and sulfofructose-1-phosphate aldolase. Our data shows that despite the superficial similarity of this pathway to glycolysis, the sulfoglycolytic enzymes are specific for SQ metabolites and are not catalytically active on related metabolites from glycolytic pathways. This observation is rationalized by 3D structures of each of these enzymes, which reveal the presence of conserved sulfonate-binding pockets. We show that SQ isomerase acts preferentially on the b-anomer of SQ and reversibly produces both SF and sulforhamnose (SR), a previously unknown sugar that acts as a transcriptional regulator for the transcriptional repressor CsqR that regulates SQ-utilisation. We also demonstrate that SF kinase is a key regulatory enzyme for the pathway that experiences complex allosteric modulation by the metabolites AMP, ADP, ATP, F6P, FBP, PEP, and citrate. This body of work provides fresh insights into the mechanism, specificity and regulation of sulfoglycolysis and has important implications for understanding how this biochemistry interfaces with central metabolism in prokaryotes to process this major repository of biogeochemical sulfur.

Structural insights into the catalytic mechanism of the yeast pyridoxal 5-phosphate synthase Snz1

2010 ◽  
Vol 432 (3) ◽  
pp. 445-454 ◽  
Author(s):  
Xuan Zhang ◽  
Yan-Bin Teng ◽  
Jian-Ping Liu ◽  
Yong-Xing He ◽  
Kang Zhou ◽  
...  
Keyword(s):  
Binding Site ◽  
Vitamin B6 ◽  
Active Sites ◽  
Biochemical Analysis ◽  
Catalytic Mechanism ◽  
De Novo ◽  
Active Form ◽  
Dihydroxyacetone Phosphate ◽  
Final Destination ◽  
Structural Insights

In most eubacteria, fungi, apicomplexa, plants and some metazoans, the active form of vitamin B6, PLP (pyridoxal 5-phosphate), is de novo synthesized from three substrates, R5P (ribose 5-phosphate), DHAP (dihydroxyacetone phosphate) and ammonia hydrolysed from glutamine by a complexed glutaminase. Of the three active sites of DXP (deoxyxylulose 5-phosphate)independent PLP synthase (Pdx1), the R5P isomerization site has been assigned, but the sites for DHAP isomerization and PLP formation remain unknown. In the present study, we present the crystal structures of yeast Pdx1/Snz1, in apo-, G3P (glyceraldehyde 3-phosphate)- and PLP-bound forms, at 2.3, 1.8 and 2.2 Å (1 Å=0.1 nm) respectively. Structural and biochemical analysis enabled us to assign the PLP-formation site, a G3P-binding site and a G3P-transfer site. We propose a putative catalytic mechanism for Pdx1/Snz1 in which R5P and DHAP are isomerized at two distinct sites and transferred along well-defined routes to a final destination for PLP synthesis.

scholarly journals Computational protein design enables a novel one-carbon assimilation pathway

2015 ◽  
Vol 112 (12) ◽  
pp. 3704-3709 ◽  
Author(s):  
Justin B. Siegel ◽  
Amanda Lee Smith ◽  
Sean Poust ◽  
Adam J. Wargacki ◽  
Arren Bar-Even ◽  
...  
Keyword(s):  
Protein Design ◽  
Biosynthetic Pathway ◽  
Carbon Fixation ◽  
Carbon Assimilation ◽  
Dihydroxyacetone Phosphate ◽  
Design Tools ◽  
Chemical Transformations ◽  
Central Metabolism ◽  
Naturally Occurring

We describe a computationally designed enzyme, formolase (FLS), which catalyzes the carboligation of three one-carbon formaldehyde molecules into one three-carbon dihydroxyacetone molecule. The existence of FLS enables the design of a new carbon fixation pathway, the formolase pathway, consisting of a small number of thermodynamically favorable chemical transformations that convert formate into a three-carbon sugar in central metabolism. The formolase pathway is predicted to use carbon more efficiently and with less backward flux than any naturally occurring one-carbon assimilation pathway. When supplemented with enzymes carrying out the other steps in the pathway, FLS converts formate into dihydroxyacetone phosphate and other central metabolites in vitro. These results demonstrate how modern protein engineering and design tools can facilitate the construction of a completely new biosynthetic pathway.

scholarly journals Structural and biochemical analysis of ATPase activity and EsxAB substrate binding of M. tuberculosis EccCb1 enzyme

2021 ◽  
Author(s):  
Arkita Bandyopadyay ◽  
Ajay Kumar Saxena
Keyword(s):  
Atpase Activity ◽  
Biochemical Analysis ◽  
Substrate Binding ◽  
Catalytic Efficiency ◽  
Size Exclusion ◽  
Enzyme Dynamics ◽  
Low Resolution ◽  
Promising Target ◽  
Binding Pockets ◽  
Resolution Structure

The EccC enzyme of M. tuberculosis ESX-1 system is a promising target for antivirulence drug development. The EccC enzyme comprises two polypeptides (i) EccCa1, a membrane bound enzyme having two ATPase domains D2 & D3 (ii) cytosolic EccCb, which contains two ATPase domains. In current study, we have analyzed the low-resolution structure of EccCb1, performed ATPase activity and EsxAB substrate binding analysis. The EccCb1 enzyme eluted as oligomer from size exclusion column and small angle X-ray scattering analysis revealed the double hexameric structure in solution. The EccCb1 enzyme showed catalytic efficiency (kcat/KM)~ 0.020 micromolar-1 min-1, however ~3.7 fold lower than its D2 and ~1.7 fold lower than D3 domains respectively. The D2 and D3 domains exhibited the ATPase activity and mutation of residues involved in ATP+Mg2+ binding have yielded 56-94% reduction in catalytic efficiency for both D2 and D3 domains. The EccCb1 binds the EsxAB substrate with KD ~ 11.4 nM via specific groove located at C-terminal region of D3 domain. ATP binding to EccCb1 enhanced the EsxAB substrate binding by ~ 3 fold, indicating ATPase energy involvement in EsxAB substrate translocation. We modeled the dodecameric EccCb1+EsxAB+ ATP+Mg2+ complex, which showed the binding pockets involved in ATP+Mg2+ and EsxAB substrate binding. The enzyme dynamics involved in ATP+Mg2+ and EsxAB substrate recognition were identified and showed the enhanced stability of EccCb1 enzyme as a result of ligand binding. Overall, our structural and biochemical analysis showed the low-resolution structure and mechanism involved in ATPase activity and EsxAB substrate binding and dynamics involved in EsxAB substrate and ATP+Mg2+ recognition. Overall, our structural and biochemical data on EccCb1 will contribute significantly in development of antivirulence inhibitors, which will prevent virulence factor secretion by M. tuberculosis ESX-1 system.

Mitochondria in Cardiomyopathy: Alterations in Size, Number and Internal Structures

1968 ◽  
Vol 26 ◽  
pp. 126-127
Author(s):  
George Hug ◽  
William K. Schubert
Keyword(s):  
Heart Failure ◽  
Biochemical Analysis ◽  
Left Ventricular ◽  
Size Number ◽  
Internal Structures ◽  
Left Ventricular Outflow ◽  
Chest X Ray ◽  
Myocardial Fibers ◽  
Muscle Biopsies ◽  
Needle Liver Biopsy

A white boy six months of age was hospitalized with respiratory distress and congestive heart failure. Control of the heart failure was achieved but marked cardiomegaly, moderate hepatomegaly, and minimal muscular weakness persisted.At birth a chest x-ray had been taken because of rapid breathing and jaundice and showed the heart to be of normal size. Clinical studies included: EKG which showed biventricular hypertrophy, needle liver biopsy which showed toxic hepatitis, and cardiac catheterization which showed no obstruction to left ventricular outflow. Liver and muscle biopsies revealed no biochemical or histological evidence of type II glycogexiosis (Pompe's disease). At thoracotomy, 14 milligrams of left ventricular muscle were removed. Total phosphorylase activity in the biopsy specimen was normal by biochemical analysis as was the degree of phosphorylase activation. By light microscopy, vacuoles and fine granules were seen in practically all myocardial fibers. The fibers were not hypertrophic. The endocardium was not thickened excluding endocardial fibroelastosis. Based on these findings, the diagnosis of idiopathic non-obstructive cardiomyopathy was made.

3H-galactose and 3H-glucose incorporation into newly synthesized hepatic glycogen in control-fed rats

1986 ◽  
Vol 44 ◽  
pp. 292-293
Author(s):  
J.E. Michaels ◽  
S.A. Garfield ◽  
J.T. Hung ◽  
S.S. Smith ◽  
R.R. Cardell
Keyword(s):  
Biochemical Analysis ◽  
Glycogen Synthesis ◽  
Hepatic Glycogen ◽  
Electron Microscopic ◽  
Once Daily ◽  
Tail Vein ◽  
Tracer Dose ◽  
Glucose Incorporation ◽  
Wet Weight

3H-galactose (gal) and 3H-glucose (glu) were compared to determine which compound was preferable for pulse labeling newly formed hepatic glycogen. Control fed rats were used to achieve substantial and consistent levels of hepatic glycogen and to stimulate glycogen synthesis.Rats fed once daily for 4 hr achieved hepatic glycogen levels > 3% wet weight liver prior to injection by tail vein of a tracer dose of 3H-gal or 3H-glu. The rats were sacrificed 15-120 min later and liver was prepared by routine techniques for light (LM) and electron microscopic (EM) radioautography (RAG) and biochemical analysis.

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