scholarly journals Redirecting Reductant Flux into Hydrogen Production via Metabolic Engineering of Fermentative Carbon Metabolism in a Cyanobacterium

2010 ◽  
Vol 76 (15) ◽  
pp. 5032-5038 ◽  
Author(s):  
Kelsey McNeely ◽  
Yu Xu ◽  
Nick Bennette ◽  
Donald A. Bryant ◽  
G. Charles Dismukes
Keyword(s):  
Metabolic Engineering ◽  
Carbon Metabolism ◽  
Anaerobic Metabolism ◽  
Lactate Production ◽  
Energy Conversion Efficiency ◽  
Liquid Chromatography Mass Spectrometry ◽  
Wild Type ◽  
Photoautotrophic Growth ◽  
Principal Source ◽  
Anaerobic Conversion

ABSTRACT Some aquatic microbial oxygenic photoautotrophs (AMOPs) make hydrogen (H2), a carbon-neutral, renewable product derived from water, in low yields during autofermentation (anaerobic metabolism) of intracellular carbohydrates previously stored during aerobic photosynthesis. We have constructed a mutant (the ldhA mutant) of the cyanobacterium Synechococcus sp. strain PCC 7002 lacking the enzyme for the NADH-dependent reduction of pyruvate to d-lactate, the major fermentative reductant sink in this AMOP. Both nuclear magnetic resonance (NMR) spectroscopy and liquid chromatography-mass spectrometry (LC-MS) metabolomic methods have shown that autofermentation by the ldhA mutant resulted in no d-lactate production and higher concentrations of excreted acetate, alanine, succinate, and hydrogen (up to 5-fold) compared to that by the wild type. The measured intracellular NAD(P)(H) concentrations demonstrated that the NAD(P)H/NAD(P)+ ratio increased appreciably during autofermentation in the ldhA strain; we propose this to be the principal source of the observed increase in H2 production via an NADH-dependent, bidirectional [NiFe] hydrogenase. Despite the elevated NAD(P)H/NAD(P)+ ratio, no decrease was found in the rate of anaerobic conversion of stored carbohydrates. The measured energy conversion efficiency (ECE) from biomass (as glucose equivalents) converted to hydrogen in the ldhA mutant is 12%. Together with the unimpaired photoautotrophic growth of the ldhA mutant, these attributes reveal that metabolic engineering is an effective strategy to enhance H2 production in AMOPs without compromising viability.

scholarly journals Sinorhizobium meliloti Mutants Lacking Phosphotransferase System Enzyme HPr or EIIA Are Altered in Diverse Processes, Including Carbon Metabolism, Cobalt Requirements, and Succinoglycan Production

2008 ◽  
Vol 190 (8) ◽  
pp. 2947-2956 ◽  
Author(s):  
Catalina Arango Pinedo ◽  
Ryan M. Bringhurst ◽  
Daniel J. Gage
Keyword(s):  
Carbon Metabolism ◽  
Sinorhizobium Meliloti ◽  
Carbon Sources ◽  
Deletion Mutants ◽  
Symbiotic Relationship ◽  
Wild Type ◽  
Phosphotransferase System ◽  
Carbon Utilization ◽  
Carbon Regulation ◽  
Extreme Sensitivity

ABSTRACT Sinorhizobium meliloti is a member of the Alphaproteobacteria that fixes nitrogen when it is in a symbiotic relationship. Genes for an incomplete phosphotransferase system (PTS) have been found in the genome of S. meliloti. The genes present code for Hpr and ManX (an EIIAMan-type enzyme). HPr and EIIA regulate carbon utilization in other bacteria. hpr and manX in-frame deletion mutants exhibited altered carbon metabolism and other phenotypes. Loss of HPr resulted in partial relief of succinate-mediated catabolite repression, extreme sensitivity to cobalt limitation, rapid die-off during stationary phase, and altered succinoglycan production. Loss of ManX decreased expression of melA-agp and lac, the operons needed for utilization of α- and β-galactosides, slowed growth on diverse carbon sources, and enhanced accumulation of high-molecular-weight succinoglycan. A strain with both hpr and manX deletions exhibited phenotypes similar to those of the strain with a single hpr deletion. Despite these strong phenotypes, deletion mutants exhibited wild-type nodulation and nitrogen fixation when they were inoculated onto Medicago sativa. The results show that HPr and ManX (EIIAMan) are involved in more than carbon regulation in S. meliloti and suggest that the phenotypes observed occur due to activity of HPr or one of its phosphorylated forms.

D-lactate production in the acorn barnacle Balanus glandula (Darwin, 1854) (Cirripedia: Balanidae) under emersion stress

2020 ◽  
Vol 40 (6) ◽  
pp. 739-745
Author(s):  
Xenia L Rangaswami ◽  
Gordon T Ober ◽  
Sarah E Gilman
Keyword(s):  
Anaerobic Metabolism ◽  
Lactate Production ◽  
Aerobic Respiration ◽  
Balanus Glandula ◽  
Air Exposure ◽  
Air Temperatures ◽  
Acorn Barnacle ◽  
The Pacific ◽  
Response To Stress ◽  
Us West

Abstract Anaerobic metabolism is an important response to stress in many organisms. Intertidal species often face heat stress during low tide. Balanus glandula (Darwin, 1854) is a high-shore intertidal barnacle common to the Pacific that experiences prolonged low-tide air exposure. It is not known whether B. glandula uses anaerobic metabolism during emersion, or if its use varies by latitude. We measured low tide D-lactate production in two US west coast populations of B. glandula separated by 14 degrees of latitude. We exposed barnacles to seven low-tide air temperatures (10, 15, 20, 25, 30, 35, and 38 °C) for which aerobic respiration has been previously measured. Our northern population of B. glandula increased D-lactate production at high air temperatures where aerobic metabolic depression is known to occur, indicating sublethal stress. In contrast, our southern population showed little increase in D-lactate over the same temperature range, coincident with high aerobic respiration across those temperatures. In a second experiment, we quantified D-lactate at 1, 2, and 6 hours post-emersion for northern B. glandula exposed to either a 10 or 38 °C low tide, to measure their potential lactate usage. While D-lactate was elevated at 38 °C compared to the 10 °C control immediately following low tide exposure, it dropped to control levels, and was likely excreted, within 1 hour of re-immersion. Our results suggest that the low latitude population of B. glandula may be more resilient to climate change than its high latitude counterpart in the absence of adaptation, which has strong implications for species distribution.

Effects of varying hematocrit on intestinal oxygen uptake in neonatal lambs

1985 ◽  
Vol 248 (4) ◽  
pp. G432-G436 ◽  
Author(s):  
I. R. Holzman ◽  
B. Tabata ◽  
D. I. Edelstone
Keyword(s):  
Blood Flow ◽  
Anaerobic Metabolism ◽  
Lactate Production ◽  
Oxygen Availability ◽  
Intestinal Blood Flow ◽  
Oxygen Extraction ◽  
Base Line ◽  
Wide Range ◽  
O2 Extraction ◽  
Neonatal Lambs

We chronically catheterized 15 newborn lambs (9.5 +/- 2.8 days) and measured intestinal blood flow (Qi) by the radionuclide microsphere technique at hematocrit levels ranging from 10 to 55%. Seven animals were made progressively anemic and eight polycythemic by means of exchange transfusions. Using the Fick principle, we calculated intestinal oxygen delivery (Di o2), oxygen consumption (Vi o2), and oxygen extraction. Initial base-line values were Qi = 195.5 ml . min-1 . 100 g intestine-1, Di o2 = 22.1 ml . min-1 . 100 g-1, Vi o2 = 4.8 ml . min-1 . 100 g-1, and O2 extraction = 22.5%. As the hematocrit was lowered, Di o2 decreased and O2 extraction increased and vice versa when the hematocrit was raised. Vi o2 remained constant, but Qi did not correlate with changes in hematocrit. However, intestinal blood flow, as a percent distribution of total blood flow, decreased with lower hematocrit levels. At no time was there any evidence of anaerobic metabolism as measured by excess lactate production. Our data indicate that the intestines of neonatal lambs are capable of maintaining their metabolic needs over a wide range of oxygen availability induced by a changing hematocrit. The primary mechanism is through alteration of oxygen extraction. Within the range of our experiments, no critically low oxygen availability was attained at which anaerobic metabolism became significant.

scholarly journals Suppressor Mutations in the Study of Photosystem I Biogenesis: sll0088 Is a Previously Unidentified Gene Involved in Reaction Center Accumulation in Synechocystis sp. Strain PCC 6803

2003 ◽  
Vol 185 (13) ◽  
pp. 3878-3887 ◽  
Author(s):  
Jianping Yu ◽  
Gaozhong Shen ◽  
Tao Wang ◽  
Donald A. Bryant ◽  
John H. Golbeck ◽  
...  
Keyword(s):  
Photosystem I ◽  
Point Mutations ◽  
Kanamycin Resistance ◽  
Negative Regulator ◽  
Binding Motif ◽  
Wild Type ◽  
Photoautotrophic Growth ◽  
Suppressor Mutations ◽  
Mutant Strains ◽  
Pcc 6803

ABSTRACT In previous work, some members of our group isolated mutant strains of Synechocystis sp. strain PCC 6803 in which point mutations had been inserted into the psaC gene to alter the cysteine residues to the FA and FB iron-sulfur clusters in the PsaC subunit of photosystem I (J. P. Yu, I. R. Vassiliev, Y. S. Jung, J. H. Golbeck, and L. McIntosh, J. Biol. Chem. 272:8032-8039, 1997). These mutant strains did not grow photoautotrophically due to suppressed levels of chlorophyll a and photosystem I. In the results described here, we show that suppressor mutations produced strains that are capable of photoautotrophic growth at moderate light intensity (20 μmol m−2 s−1). Two separate suppressor strains of C14SPsaC, termed C14SPsaC-R62 and C14SPsaC-R18, were studied and found to have mutations in a previously uncharacterized open reading frame of the Synechocystis sp. strain PCC 6803 genome named sll0088. C14SPsaC-R62 was found to substitute Pro for Arg at residue 161 as the result of a G482→C change in sll0088, and C14SPsaC-R18 was found to have a three-amino-acid insertion of Gly-Tyr-Phe following Cys231 as the result of a TGGTTATTT duplication at T690 in sll0088. These suppressor strains showed near-wild-type levels of chlorophyll a and photosystem I, yet the serine oxygen ligand to FB was retained as shown by the retention of the S ≥ 3/2 spin state of the [4Fe-4S] cluster. The inactivation of sll0088 by insertion of a kanamycin resistance cartridge in the primary C14SPsaC mutant produced an engineered suppressor strain capable of photoautotrophic growth. There was no difference in psaC gene expression or in the amount of PsaC protein assembled in thylakoids between the wild type and an sll0088 deletion mutant. The sll0088 gene encodes a protein predicted to be a transcriptional regulator with sequence similarities to transcription factors in other prokaryotic and eukaryotic organisms, including Arabidopsis thaliana. The protein contains a typical helix-turn-helix DNA-binding motif and can be classified as a negative regulator by phylogenetic analysis. This suggests that the product of sll0088 has a role in regulating the biogenesis of photosystem I.

Glucose uptake by diaphragms from rats subjected to hemorrhagic shock

1964 ◽  
Vol 206 (2) ◽  
pp. 317-320 ◽  
Author(s):  
William R. Drucker ◽  
John C. DeKiewiet
Keyword(s):  
Skeletal Muscle ◽  
Hemorrhagic Shock ◽  
Glucose Uptake ◽  
Anaerobic Metabolism ◽  
Lactate Production ◽  
Bicarbonate Buffer ◽  
Metabolic Alterations ◽  
Shock Models ◽  
The Media ◽  
Intracellular Energy

The marked metabolic alterations that occur in hemorrhagic shock have been ascribed to tissue anoxia occasioned by hypovolemia. Other investigators, utilizing different shock models, have explained the initial metabolic changes as secondary to humoral changes. In skeletal muscle, anoxia is known to cause an increased glucose uptake, whereas epinephrine causes a decreased uptake. The present work was undertaken to explore some alterations in carbohydrate metabolism during hemorrhagic shock in rats, when both tissue anoxia and an altered humoral state are present. Hemidiaphragms from rats subjected to a standardized hemorrhagic shock procedure and from control rats were excised and incubated aerobically in bicarbonate buffer containing glucose. After 1 hr of incubation aliquots of the media were analyzed for glucose and lactate. The results demonstrated a significantly greater glucose uptake and lactate production by the diaphragms from the bled rats. The data suggest that, during hemorrhagic shock in rats, tissue anoxia leads to a predominance of anaerobic metabolism and a severe depletion of intracellular energy, resulting in an increased uptake of glucose in skeletal muscle despite the concomitant altered humoral state which ordinarily would inhibit glucose uptake.

Improvement of hydrogen production by metabolic engineering of Escherichia coli: Modification on both the PTS system and central carbon metabolism

2020 ◽  
Vol 45 (9) ◽  
pp. 5687-5696 ◽  
Author(s):  
Victor E. Balderas-Hernandez ◽  
Kathya P. Landeros Maldonado ◽  
Arturo Sánchez ◽  
Adam Smoliński ◽  
Antonio De Leon Rodriguez
Keyword(s):  
Escherichia Coli ◽  
Metabolic Engineering ◽  
Hydrogen Production ◽  
Carbon Metabolism ◽  
Central Carbon Metabolism ◽  
Central Carbon

Aerobic and anaerobic contributions to energy metabolism in perfused isolated sea raven (Hemitripterus americanus) hearts

1983 ◽  
Vol 61 (8) ◽  
pp. 1880-1883 ◽  
Author(s):  
William R. Driedzic ◽  
Donna L. Scott ◽  
Anthony P. Farrell
Keyword(s):  
Anaerobic Metabolism ◽  
Lactate Production ◽  
Fish Heart ◽  
Atp Production ◽  
Isolated Hearts ◽  
Relative Contribution ◽  
Sea Raven ◽  
Hemitripterus Americanus ◽  
Aerobic And Anaerobic ◽  
Experimental Preparation

The relative contribution of aerobic and anaerobic metabolism to ATP production was assessed in sea raven (Hemitripterus americanus) hearts. The problem was approached by measuring the rates of oxygen consumption and lactate production by perfused isolated hearts performing mechanical work. In the experimental preparation aerobic metabolism could account for essentially all of the ATP synthesized; as such, the organization of metabolism in this fish heart appears similar to reptilian and mammalian hearts under conditions of adequate oxygen availability.

Modelling the peroxisomal carbon leak during lipid mobilization in Arabidopsis

2010 ◽  
Vol 38 (5) ◽  
pp. 1230-1233 ◽  
Author(s):  
Mark A. Hooks ◽  
Elizabeth Allen ◽  
Jonathan A.D. Wattis
Keyword(s):  
Carbon Metabolism ◽  
Glyoxylate Cycle ◽  
Seedling Development ◽  
Wild Type ◽  
Acetyl Coa ◽  
Primary Metabolite ◽  
Lipid Mobilization ◽  
Acetate Assimilation ◽  
Development Levels ◽  
The Glyoxylate Cycle

Mutation of the ACN1 (acetate non-utilizing 1) locus of Arabidopsis results in altered acetate assimilation into gluconeogenic sugars and anapleurotic amino acids and leads to an overall depression in primary metabolite levels by approx. 50% during seedling development. Levels of acetyl-CoA were higher in acn1 compared with wild-type, which is counterintuitive to the activity of ACN1 as a peroxisomal acetyl-CoA synthetase. We hypothesize that ACN1 recycles free acetate to acetyl-CoA within peroxisomes in order that carbon remains fed into the glyoxylate cycle. When ACN1 is not present, carbon in the form of acetate can leak out of peroxisomes and is reactivated to acetyl-CoA within the cytosol. Kinetic models incorporating estimates of carbon input and pathway dynamics from a variety of literature sources have proven useful in explaining how ACN1 may prevent the carbon leak and even contribute to the control of peroxisomal carbon metabolism.

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