Metabolomics/Metabolites/Carbohydrates/Glucose
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Glycomics and Glycoproteomics
editThe fields of glycomics and glycoproteomics involve the study of a cell, tissue, or organism's glycan and glycoprotein content. Among the numerous bioanalytical techniques utilized in these fields are 2D-PAGE, MALDI-TOFMS, LC-QMS, LC-ICRMS, LC-QTOF, and LC-IMS-TOF.
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Glycan
Otherwise known as a polysaccharide, a glycan is a polymer of monosaccharide units joined by glycosidic bonds.
Glycoprotein
Glycoproteins are proteins with covalently attached carbohydrates . These carbohydrates can be bonded via the OH group of serine or threonine, resulting in O glycosidic bonds. Carbohydrates may also be bonded via the amide group of asparagine, which results in an N glycosidic bond.
Glycome
The glycome represents the total set of glycans present in a cell or an organism.
Glycosylation
Technology and Analytical Techniques
A ratiometric lectin microarray approach was employed to analyze the dynamics of the mammalian glycome. In this experiment, the microarray approach was utilized to analyze the rapid evaluation of differences in the glycosylation of heterogeneous mammalian samples. This research is indicative of the significant improvements that have been, and will be made in the field of glycomics. Further research and development of the microarray approach to analyzing glycans is predicted to allow for significant insight into how glycans encode biological information in complex systems.
The approach employed in this experiment, comprising of a biological referencing system coupled to the ratiometric approach, allowed for a sensitive analytical method that enabled researchers to detect the subtle differences between samples. This analytical system has evidently provided reproducible, semiquantitative lectin-binding analyses of complex cell-surface glycosylation, and already been applied to the analysis of blood cell differentiation. By allowing for this simple high-throughput method to obtain an overall view of N- and O-linked glycosylation, this technology has made immeasurable improvement upon past methods of glycomic analysis. In addition, the ratiometric microarray has the potential to obtain an overall view of glycolipid content, at an intermediate level of substructural resolution. Microarray technology may also be used to verify the results from other bioanalytical techniques, such as RT-PCR. Future prospects for the microarray comprise of expanding the array to include antibodies, new more highly selective lectins, as well as other glycan binding proteins. This expansion will allow researchers to gather more detailed information about the mammalian glycome. Fluctuation of glycan content associated with dynamic biological states, including disease states, may also be more systematically identified, as this emerging technology advances. Furthermore, the microarray is expected to reveal direct linkages between the mammalian glycome and genome.
References:
Indiana University National Center for Glycomics and Glycoproteomics
{Text Book}
A ratiometric lectin microarray approach to analysis of the dynamic mammalian glycome
KEGG Pathway
editNote: These could not be regularly hyperlinked as for some reason they set off wikibook's spam filters.
Glycolysis / Gluconeogenesis
http://www.genome.ad.jp/dbget-bin/show_pathway?map00010+C00031
Pentose phosphate pathway
http://www.genome.ad.jp/dbget-bin/show_pathway?map00030+C00031
Galactose metabolism
http://www.genome.ad.jp/dbget-bin/show_pathway?map00052+C00031
Starch and sucrose metabolism
http://www.genome.ad.jp/dbget-bin/show_pathway?map00500+C00031
Streptomycin biosynthesis
http://www.genome.ad.jp/dbget-bin/show_pathway?map00521+C00031
Indole and ipecac alkaloid biosynthesis
http://www.genome.ad.jp/dbget-bin/show_pathway?map00901+C00031
ABC transporters – General
http://www.genome.ad.jp/dbget-bin/show_pathway?map02010+C00031
Two-component system – General
http://www.genome.ad.jp/dbget-bin/show_pathway?map02020+C00031
Insulin signaling pathway
http://www.genome.ad.jp/dbget-bin/show_pathway?map04910+C00031
Type II diabetes mellitus
http://www.genome.ad.jp/dbget-bin/show_pathway?map04930+C00031
MetaCyc Pathways
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Web Resources
Wikipedia: Glucose
http://en.wikipedia.org/wiki/Glucose
This is the Wikipedia entry for glucose. It presents a broad overview of glucose including it physical and chemical properties, chemical structures and isomers, natural and commercial sources of production and roles it plays in cellular metabolism. The site also presents the glycolysis pathway with links to all enzymes and intermediates involved.
New terms:
Chiral – Structural characteristic of a molecule that makes it impossible to superimpose it on its mirror image.
Anomer – A cyclic stereoisomer whose sole conformational difference involves the arrangement of atoms or groups in the aldehyde or ketone group.
Fischer projection – A two-dimensional representation of a three-dimensional organic molecule.
Haworth projection – A way of representing the cyclic structure of monosaccharides with a simple three-dimensional perspective.
Mutarotation – A change in the optical rotation reducing sugars.
Rotamers – One of a set of conformers arising from restricted rotation about one single bond.
Abiotic – Non-living chemical and physical factors of the environment.
Erythrocyte – Red blood cell.
This site provides background information on the glucose molecule itself as well as a lot of concepts discussed in class such as glycolysis, citric acid cycle, glycogen, protein and lipid synthesis etc. The outline of the glycolysis metabolic pathway with links to all enzymes and intermediates involved would be an especially useful aid in its study.
MedBio
This site provides information on carbohydrate structure and function, glucose metabolism and control and diabetes and other related metabolic disorders. The site serves as a great resource because it not only provides information on standalone topics, but contains a large amount or articles and explanations that integrate glucose metabolism into broader concepts that go above and beyond what was covered in class in a manner that is easier to understand than the textbook. The site also has summaries, explanations and links to other medical textbooks and websites on the topic.
New terms:
Glycemic – The presence of glucose in the blood.
Hepatic – Relating to the liver.
Renal – Relating to the kidney.
Lipolysis – The hydrolysis of fats into fatty acids and glycerol.
Adsorbtion – The gathering of a substance on the surface of a layer.
Biphasic - Having two distinct phases.
All concepts discussed in class are covered in this site, including, but not limited to, glucose structure and function, glycolysis, gluconeogenesis, the citric acid cycle, energy storage and metabolic pathways of synthesis and degradation that use glucose as a metabolite.
Glucose Tolerance Test
http://www.labtestsonline.org/understanding/analytes/glucose/test.html
This site describes the nature of the glucose tolerance test in the clinical diagnosis of diabetes. It explains the nature of the disease, and the consequences of hyper and hypoglycemia. It thoroughly explains the process and the meaning of the results, listing detailed information on how blood glucose levels change in a diabetic compared to a healthy individual.
New terms:
Hypoglycemia – An abnormally low level of glucose in the blood.
Hyperglycemia – An abnormally high level of glucose in the blood.
Gestational diabetes – A condition in which women without previously diagnosed diabetes exhibit high blood glucose levels during pregnancy. It is believed that the hormones produced during pregnancy reduce a woman's sensitivity to insulin, resulting in high blood sugar levels.
Type I diabetes – An autoimmune disease that results in the permanent destruction of insulin producing beta cells of the pancreas.
Type II diabetes - A metabolic disorder that is primarily characterized by insulin resistance, relative insulin deficiency and hyperglycemia.
This site provides a good launching off point for a more detailed examination of our brief discussion of diabetes in class. It explains the clinical symptoms of diabetes as well as offering a discussion and links to other medical conditions that affect blood glucose and insulin levels.
Peer-reviewed articles
The Role of Glucose Metabolites in the Activation and Translocation of Glycogen Synthase by Insulin in 3T3-L1 Adipocytes
http://www.jbc.org/cgi/content/full/274/39/27497
This study investigated the effects of differing concentrations of various glucose metabolites on the action of glycogen synthase (the enzyme responsible for glycogen synthesis) in mouse adiposite tissue. Its main findings were that increasing the levels of insulin, glucose-6-phosphate and glucosamine-6-phosphate increased the levels if glycogen synthase activity. Conversely, decreasing the levels of available glucose in the cells decreased the activity of glycogen synthase.
New terms:
Glycogenolytic – The biochemical breakdown of glycogen to glucose.
Immunoblot – A laboratory procedure in which proteins that have been separated by electrophoresis are transferred onto nitrocellulose sheets and are identified by their reaction with labeled antibodies.
Isoform – Any of two or more functionally similar proteins that have a similar but not identical amino acid sequence and are either encoded by different genes or by RNA transcripts from the same gene which have had different exons removed.
Endogenous – Pertaining to the metabolism of nitrogenous elements of cells and tissues.
Lysate – The mixture of substances formed by the lysis of cells.
Supernatant – Floating above or on the surface.
Ultracentrifugation – A high-velocity centrifuge used in the separation of colloidal or submicroscopic particles.
Subcellular – Contained within a cell.
As we have seen throughout the course, the regulation of metabolic processes involves the interaction amongst many different integrated metabolic pathways and substances, and the study clearly illustrates this concept. It shows how cellular metabolites have many different roles, with the substrates or products of one pathway actually acting as regulators of another pathway and also illustrates different methods of enzyme regulation such as translocation, sequestration and allosteric control.
Determination of glucose metabolites in stored erythrocytes and in erythrocytes from patients with thalassemia by analytical isotachophoresis
This study used an isotachophoretic method to study the concentrations of all the metabolites of glycolysis in erythrocytes (red blood cells). This is relevant since red blood cells (RBCs) rely on glycolysis as their main source of ATP production. RBCs from healthy individuals as well as those suffering from thalassemia were used in the study. It was found that concentrations of 2,3-bisphosphoglycerate were elevated in thalassemia RBCs compared to those from healthy individuals.
New terms:
Isotachophoresis – A technique in analytical chemistry used to separate charged particles using a discontinuous electrical field to create sharp boundaries between the sample constituents.
Thalassemia – Hereditary disease causing faulty haemoglobin synthesis.
Electrolyte – Any inorganic compounds, mainly sodium, potassium, magnesium, calcium, chloride, and bicarbonate, that dissociate in biological fluids into ions capable of conducting electrical currents and constituting a major force in controlling fluid balance within the body.
Electrophoresis – A method of separating substances, especially proteins, and analyzing molecular structure based on the rate of movement of each component in a colloidal suspension while under the influence of an electric field.
Anaemia – A quantitative deficiency of the hemoglobin, often accompanied by a reduced number of red blood cells and causing pallor, weakness, and breathlessness.
Microcytosis – A blood disorder characterized by the presence of microcytes (abnormally small red blood cells) in the blood; often associated with anemia.
Hypochromia – An anaemic condition due to a deficiency of haemoglobin in the red blood cells causing a lack of colour or pigmentation.
This study featured an in depth analysis of the glycolysis pathway, describing an experimental method that can be used to analyse and determine the concentration of glycolysis metabolites. This information would be an excellent supplement to our class study of the glycolysis pathway in terms of proving background information on how the various intermediate metabolites in the pathway can be detected and measured in the first place. It also illustrates another example of the concept of an inborn error of metabolism causing the accumulation of intermediate metabolites in a pathway.
Glucose Metabolites Inhibit Protein Phosphatases and Directly Promote Insulin Exocytosis in Pancreatic ß-Cells
http://endo.endojournals.org/cgi/content/full/endo;143/12/4592
This study examined the causes of insulin excretion from mice pancreatic β-cells. It found that metabolites of glycolysis and the citric acid cycle such as fructure-1,6-bisphosphate, phosphoenolpyruvate, 3-phosphoglycerate, citrate and oxaloacetate significantly enhance insulin exocytosis from pancreatic β-cells by inactivating ser/thr phosphatases.
New terms: Permeabilized – Made permeable.
Homogenize - to form by a consistent solution by blending unlike elements.
Normoglycemia – The presence of a normal concentration of glucose in the blood.
Anaplerosis – Reactions that form intermediates of the TCA or citric acid cycle.
Mitogenesis – The induction of mitosis.
Secretagogue – A substance or situation that promotes secretion.
This article again further expands on our discussions of diabetes, glycolysis and the citric acid cycle. It clearly illustrates the concept that metabolic pathways are not simple and linear as may seem when they are presented on a one-by-one in the textbook, but are in fact intrinsically linked and interconnected in terms of locations, metabolites and regulatory enzymes. Finally, since this paper is relatively new, it also shows that we a far from having a complete holistic understanding of cellular metabolism processes and are constantly learning new functions of ‘old’ molecules.