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Garth L. Fletcher - One of the best experts on this subject based on the ideXlab platform.
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Evolutionary relationships between AFP-producing fishes and the similarities between type I AFPs.
2013Co-Authors: Laurie A. Graham, Garth L. Fletcher, Rod S. Hobbs, Peter L. DaviesAbstract:A) Phylogenetic relationships (not to scale) among AFP-producing fish from analysis of complete mitochondrial genomes [67]–[69] or selected nuclear and mitochondrial sequences [45]. Estimated divergence times (Ma, some with 95% highest posterior density limits) [45]–[47] are shown at some nodes. Species names are colored by AF(G)P type as indicated on the right. Representative ribbon structures are shown for types II, III, and I AFPs (PDB 2PY2, 1HG7, 1WFA from top to bottom, red = helix, green = strand, gray = coil). The colored bars at the bottom indicate climate differences marked by the presence (blue) or absence (red) of large ice sheets. Common names of representative AFP-producing fish are indicated but their scientific names are as follows; herring (Clupea harengus), Arctic cod (Boreogadus saida), cunner (Tautogolabrus adspersus), ocean pout (Zoarces americanus), Atlantic snailfish (Liparis atlanticus), dusky snailfish (Liparis gibbus), sea raven (Hemitripterus americanus), longhorn sculpin (Myoxocephalus octodecemspinosus), shorthorn sculpin (Myoxocephalus scorpius), Antarctic toothfish (Dissostichus mawsoni), winter flounder (Pseudopleuronectes americanus) and rainbow smelt (Osmerus mordax). B) Alignment of representative type I skin AFPs from three fishes from three separate orders (winter flounder (M63478.1), longhorn sculpin (AF306348.1) and cunner (JF937681.2). Potential or known ice-binding residues within the 11-aa repeat that show an i, i+4, i+8 spacing pattern are indicated with plus symbols (Ala) and number symbols (Thr) with asterisks denoting residues that are identical in all sequences. Acidic and basic residues are in red and blue font respectively, with Ala highlighted yellow and Thr in white font with black highlighting. Potential helix-stabilizing salt bridges consisting of basic and acidic residues with the more effective i, i+4 separation [70] are double underlined. The cunner isoform is also found in blood [42].
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Isolation and purification of antifreeze proteins from skin tissues of snailfish, cunner and sea raven.
Biochimica et biophysica acta, 2004Co-Authors: Robert P. Evans, Garth L. FletcherAbstract:Antifreeze proteins/polypeptides (AFPs), which are found in diverse species of marine fish, are grouped into four distinct classes (types I-IV). The discovery of skin-specific type I AFPs established that this class contains distinct isoforms, liver-type and skin-type, which are encoded by separate gene families. In this study, type I AFPs were isolated and partially characterized from skin tissues of Atlantic snailfish (Liparis atlanticus) and cunner (Tautogolabrus adspersus). Interestingly, evidence from this study indicates that snailfish type I AFPs synthesized in skin tissues are identical to those circulating in their blood plasma. Furthermore, type II AFPs that are identical to those expressed in liver for export into blood were purified from sea raven (Hemitripterus americanus) skin tissue extracts. It is clear that epithelial tissues are an important source for antifreeze expression to enhance the complement of AFPs that protect fish from freezing in extreme cold environments. In addition, the evidence generated in this study demonstrates that expression of AFPs in fish skin is a widespread phenomenon that is not limited to type I proteins.
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Tissue distribution of fish antifreeze protein mRNAs
Canadian Journal of Zoology, 1992Co-Authors: Zhiyuan Gong, Garth L. Fletcher, Choy L. HewAbstract:The presence of fish antifreeze protein (AFP) mRNA was examined in a variety of tissues from the winter flounder (Pseudopleuronectes americanus), sea raven (Hemitripterus americanus), and ocean pou...
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Structural and functional similarity between fish antifreeze proteins and calcium-dependent lectins.
Biochemical and Biophysical Research Communications, 1992Co-Authors: K. Vanya Ewart, Boris Rubinsky, Garth L. FletcherAbstract:A cDNA for a type II antifreeze protein was isolated from liver of smelt (Osmerus mordax). The predicted protein sequence is homologous to that from sea raven (Hemitripterus americanus) and both show homology to a family of calcium-dependent lectins. Smelt and sea raven belong to taxonomic orders believed to have diverged prior to Cenozoic glaciation. Thus, type II antifreeze proteins appear to have evolved independently in these fish species from pre-existing calcium-dependent lectins. Sequence alignment of the antifreezes and the lectins suggest that these proteins adopt a similar fold, that the sea raven antifreeze has lost its Ca2+ binding sites, and the smelt antifreeze has retained one site. Experiments show that smelt antifreeze protein activity is responsive to Ca2+ but that of sea raven antifreeze protein is not. These results suggest that the type II fish antifreeze proteins and calcium-dependent lectins share a common ancestry, related folding structures, and functional similarity.
William R. Driedzic - One of the best experts on this subject based on the ideXlab platform.
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Oxygen consumption in myoglobin-rich and myoglobin-poor isolated fish cardiomyocytes.
The Journal of experimental zoology, 1998Co-Authors: Nicola J. N. Legate, John R. Bailey, William R. DriedzicAbstract:The function of myoglobin at the cellular level was investigated by comparing O2 consumption in isolated myoglobin-rich cardiac myocytes from the sea raven (Hemitripterus americanus) and myoglobin-poor myocytes from the ocean pout (Macrozoarces americanus). O2 consumption by sea raven myocytes, 0.21 +/- 0.04 microM O2/10(6) cells.min-1, was significantly higher than O2 consumption by ocean pout myocytes, 0.10 +/- 0.07 microM O2/10(6) cells.min-1 at high PO2. O2 consumption in sea raven myocytes treated with sodium nitrite was not significantly different than that in untreated myocytes at high PO2, but it was significantly lower than controls at low PO2. O2 consumption of sea raven myocytes treated with the mitochondrial uncoupler CCCP was not significantly different from that of control myocytes at high PO2, but it was significantly greater than untreated controls at low PO2. In ocean pout preparations, O2 consumption by nitrite-treated myocytes was significantly higher than that of untreated myocytes at high PO2, but it was not different from that of controls at low PO2. CCCP-treated ocean pout myocytes had a significantly higher oxygen consumption than that of untreated myocytes at high PO2, but oxygen consumption was not different from that of controls at low PO2. The CCCP-activated O2 consumption at low PO2 was myoglobin-dependent in that CCCP alone resulted in a threefold increase in sea raven cells over controls but had no impact on sea raven cells in the presence of nitrite or ocean pout cells treated with CCCP alone. This study further supports the contention that myoglobin only plays an important role in oxygen metabolism at low extracellular PO2's.
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Glucose metabolism by sea raven (Hemitripterus americanus) and rainbow trout (Oncorhynchus mykiss) erythrocytes
The Journal of experimental biology, 1994Co-Authors: Dawn H. Sephton, William R. DriedzicAbstract:The fate of extracellular glucose in blood isolated from sea raven (Hemitripterus americanus) and rainbow trout (Onchorhynchus mykiss) was determined. In blood from both species incubated in vitro at low physiological pH, the decrease in plasma glucose concentration was more than adequate to support oxygen consumption. Glucose disappearance could not be accounted for by increases in lactate, red blood cell (RBC) glucose or RBC glycogen concentrations. Rates of 14CO2 production from [6-14C]glucose over a 2 h incubation period were less than 1 % of metabolic rate. Only small amounts of label appeared in RBC protein, lipid or glycogen fractions relative to metabolic rates, but label accumulated in the intracellular acid-soluble fraction (presumably free glucose, glycolytic intermediates, amino acids, citric acid cycle intermediates, etc.) at rates consistent with oxygen consumption and glucose disappearance. The simplest explanation for the mismatch between 14CO2 production and the other estimates of metabolic rate is that incubation times were too short for equilibration to occur. A consequence is that studies of this nature cannot use 14CO2 production to elucidate rates of aerobic fuel utilization. By default, the data imply that glucose serves as a primary aerobic metabolic fuel for the RBCs, at least under some conditions.
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Metabolic Enzyme Activities, Oxygen Consumption and Glucose Utilization in Sea Raven (Hemitripterus americanus) Erythrocytes
The Journal of Experimental Biology, 1991Co-Authors: Dawn H. Sephton, Wanda Lee Macphee, William R. DriedzicAbstract:The organization of energy metabolism was assessed in erythrocytes of the sea raven ( Hemitripterus americanus ). Cell suspensions displayed a potential for aerobic glycolysis, shown by the presence of the glycolytic enzymes phosphofructokinase, aldolase and pyruvate kinase and the mitochondrial markers citrate synthase and malate dehydrogenase. Rates of oxygen uptake ( M o o2 ) and glucose uptake M glucose (as assessed by the disappearance of glucose) are closely matched in whole blood and resuspended erythrocyte preparations. Lactate does not accumulate under aerobic conditions. The enzymatic potential is well in excess of maximal rates of carbon flux in intact cells. Overall, the data reveal that sea raven erythrocytes have an aerobic metabolism that is fuelled by exogenous glucose. Calculated rates of glucose oxidation from [6- 14 C]glucose were 1000-fold lower than rates measured directly from glucose disappearance, implying that exogenous glucose is highly diluted or mixed into the intracellular pools prior to entry into the citric acid cycle.
Peter L. Davies - One of the best experts on this subject based on the ideXlab platform.
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Evolutionary relationships between AFP-producing fishes and the similarities between type I AFPs.
2013Co-Authors: Laurie A. Graham, Garth L. Fletcher, Rod S. Hobbs, Peter L. DaviesAbstract:A) Phylogenetic relationships (not to scale) among AFP-producing fish from analysis of complete mitochondrial genomes [67]–[69] or selected nuclear and mitochondrial sequences [45]. Estimated divergence times (Ma, some with 95% highest posterior density limits) [45]–[47] are shown at some nodes. Species names are colored by AF(G)P type as indicated on the right. Representative ribbon structures are shown for types II, III, and I AFPs (PDB 2PY2, 1HG7, 1WFA from top to bottom, red = helix, green = strand, gray = coil). The colored bars at the bottom indicate climate differences marked by the presence (blue) or absence (red) of large ice sheets. Common names of representative AFP-producing fish are indicated but their scientific names are as follows; herring (Clupea harengus), Arctic cod (Boreogadus saida), cunner (Tautogolabrus adspersus), ocean pout (Zoarces americanus), Atlantic snailfish (Liparis atlanticus), dusky snailfish (Liparis gibbus), sea raven (Hemitripterus americanus), longhorn sculpin (Myoxocephalus octodecemspinosus), shorthorn sculpin (Myoxocephalus scorpius), Antarctic toothfish (Dissostichus mawsoni), winter flounder (Pseudopleuronectes americanus) and rainbow smelt (Osmerus mordax). B) Alignment of representative type I skin AFPs from three fishes from three separate orders (winter flounder (M63478.1), longhorn sculpin (AF306348.1) and cunner (JF937681.2). Potential or known ice-binding residues within the 11-aa repeat that show an i, i+4, i+8 spacing pattern are indicated with plus symbols (Ala) and number symbols (Thr) with asterisks denoting residues that are identical in all sequences. Acidic and basic residues are in red and blue font respectively, with Ala highlighted yellow and Thr in white font with black highlighting. Potential helix-stabilizing salt bridges consisting of basic and acidic residues with the more effective i, i+4 separation [70] are double underlined. The cunner isoform is also found in blood [42].
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Evidence for a proprotein intermediate during maturation of type II antifreeze protein in sea raven, Hemitripterus americanus.
Biochimica et biophysica acta, 1996Co-Authors: Bernard P. Duncker, Sherry Y. Gauthier, Peter L. DaviesAbstract:The circulating Type II antifreeze protein (AFP) in sea raven is 129 amino acids (aa) long (14 kDa) and is derived from an initial 163 aa translation product that is synthesised in the liver. Signal peptide cleavage algorithms, as well as transgenic expression studies in fall armyworm cells, predict the formation of a 146 aa (16 kDa) proprotein intermediate. A protein of this size that cross-reacted with anti-sea raven AFP antibody was detected in sea raven serum using phosphate/urea SDS-PAGE, and was purified by size-exclusion chromatography and reverse-phase HPLC. N-terminal sequencing and mass spectrometry identified the protein as the predicted proAFP, and immunoblotting suggested that it is the predominant form present in liver. These results are consistent with production and storage of a proAFP intermediate in the liver, and its subsequent processing to mature AFP during or soon after its release into the circulation.
Dawn H. Sephton - One of the best experts on this subject based on the ideXlab platform.
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Glucose metabolism by sea raven (Hemitripterus americanus) and rainbow trout (Oncorhynchus mykiss) erythrocytes
The Journal of experimental biology, 1994Co-Authors: Dawn H. Sephton, William R. DriedzicAbstract:The fate of extracellular glucose in blood isolated from sea raven (Hemitripterus americanus) and rainbow trout (Onchorhynchus mykiss) was determined. In blood from both species incubated in vitro at low physiological pH, the decrease in plasma glucose concentration was more than adequate to support oxygen consumption. Glucose disappearance could not be accounted for by increases in lactate, red blood cell (RBC) glucose or RBC glycogen concentrations. Rates of 14CO2 production from [6-14C]glucose over a 2 h incubation period were less than 1 % of metabolic rate. Only small amounts of label appeared in RBC protein, lipid or glycogen fractions relative to metabolic rates, but label accumulated in the intracellular acid-soluble fraction (presumably free glucose, glycolytic intermediates, amino acids, citric acid cycle intermediates, etc.) at rates consistent with oxygen consumption and glucose disappearance. The simplest explanation for the mismatch between 14CO2 production and the other estimates of metabolic rate is that incubation times were too short for equilibration to occur. A consequence is that studies of this nature cannot use 14CO2 production to elucidate rates of aerobic fuel utilization. By default, the data imply that glucose serves as a primary aerobic metabolic fuel for the RBCs, at least under some conditions.
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Metabolic Enzyme Activities, Oxygen Consumption and Glucose Utilization in Sea Raven (Hemitripterus americanus) Erythrocytes
The Journal of Experimental Biology, 1991Co-Authors: Dawn H. Sephton, Wanda Lee Macphee, William R. DriedzicAbstract:The organization of energy metabolism was assessed in erythrocytes of the sea raven ( Hemitripterus americanus ). Cell suspensions displayed a potential for aerobic glycolysis, shown by the presence of the glycolytic enzymes phosphofructokinase, aldolase and pyruvate kinase and the mitochondrial markers citrate synthase and malate dehydrogenase. Rates of oxygen uptake ( M o o2 ) and glucose uptake M glucose (as assessed by the disappearance of glucose) are closely matched in whole blood and resuspended erythrocyte preparations. Lactate does not accumulate under aerobic conditions. The enzymatic potential is well in excess of maximal rates of carbon flux in intact cells. Overall, the data reveal that sea raven erythrocytes have an aerobic metabolism that is fuelled by exogenous glucose. Calculated rates of glucose oxidation from [6- 14 C]glucose were 1000-fold lower than rates measured directly from glucose disappearance, implying that exogenous glucose is highly diluted or mixed into the intracellular pools prior to entry into the citric acid cycle.
Bernard P. Duncker - One of the best experts on this subject based on the ideXlab platform.
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Evidence for a proprotein intermediate during maturation of type II antifreeze protein in sea raven, Hemitripterus americanus.
Biochimica et biophysica acta, 1996Co-Authors: Bernard P. Duncker, Sherry Y. Gauthier, Peter L. DaviesAbstract:The circulating Type II antifreeze protein (AFP) in sea raven is 129 amino acids (aa) long (14 kDa) and is derived from an initial 163 aa translation product that is synthesised in the liver. Signal peptide cleavage algorithms, as well as transgenic expression studies in fall armyworm cells, predict the formation of a 146 aa (16 kDa) proprotein intermediate. A protein of this size that cross-reacted with anti-sea raven AFP antibody was detected in sea raven serum using phosphate/urea SDS-PAGE, and was purified by size-exclusion chromatography and reverse-phase HPLC. N-terminal sequencing and mass spectrometry identified the protein as the predicted proAFP, and immunoblotting suggested that it is the predominant form present in liver. These results are consistent with production and storage of a proAFP intermediate in the liver, and its subsequent processing to mature AFP during or soon after its release into the circulation.