00061 - Biochemistry

Course Unit Page

Academic Year 2017/2018

Learning outcomes

At the end of the course, the student becomes familar with essential biochemical concepts: macromolecular structures (sugars, proteins, lipids, nucleic acids) and functions, catalysis, intermediary metabolism and regulation, cell signalling. Special emphasis will be given to the major experimental metodologies in biochemistry.

Course contents

Lectures

Aminoacid and proteins: structure and properties of amino acids; structure, stability and formation of the peptide bond. Weak interactions in an aqueous environment: charge-charge interactions, the hydrogen bond, dipole-dipole interactions, hydrophobic and van der Waals interactions. Primary, secondary, tertiary and quaternary structure of proteins; introduction to prediction of protein structure.

Lipids and membranes: fatty acids, tryglycerids, phospholipids and sphingolipids, cholesterol. Structure, composition, asimmetry and properties of biological membranes, membrane proteins: bacteriorhodopsin and porin. Lipids rafts.

How to explore  proteins: the proteome, protein purifiation: subcellular fractionaction, centrifugation, cromatographyc techniques, electrophoresis (SDS and native), western blot, immunofluorescence, analysis by X-ray cristallography. Bioinformatics and prediction of protein structure.

Myoglobin and hemoglobin: structure of heme, myiglobin and hemoglobin, oxygen binding, saturation curves, effect of pH and 2,3-DPG.

Enzymes: proteins as molecular catalysts, chemical reaction rates and the effects of catalysts, transition states, the  enzyme-substrate complex, some general catalytic mechanisms. Introduction to kinetics of enzymatic catalysis: the Michaelis-Menten analysis, the significance of Km, Vmax and Kcat, rearrangement of Michaelis-Menten equation (Lineweaver-Burk  plot). Enzyme inhibition: reversible and irreversible inhibition, competitive, non competitive and acompetitive inhibitors. Multisubstrate reactions. Regulation of enzyme activity: allosteric regulation (aspartate transcabamylase (ATCase), covalent modifications: reversible (phosphorylation, acetylation) and irreversible (proteolysis), regulation by control of enzymatic synthesis and degradation. Examples of some catalytic strategies: chymotrypsin and other proteases.

Signal transduction pathways: signalling strategies, first messengers, intracellular and membrane receptors. Membrane receptors types: 7TM receptors: rhodopsin and beta-adrenergic receptor, G proteins: activation/deactivation, effector systems: adenylate cyclase and phospholipase C. Receptor with thyrosine kinase activity and SH2 domain-mediated signalling,  Grb2, Ras and MAP kinases. Insulin and insulin receptor signalling: IRS1, PI-3K, PDK1, Akt and Gsk.

Introduction to metabolism: an overview of major metabolic pathways, existence of separate biosynthetic and degradative pathways, energy transductions, the ATP molecule, coenzymes and vitamines.

Carbohydrate metabolism: the molecular structure and behaviour of carbohydrates. Glycolysis, Destiny of pyruvate: fermentations, regulation of glycolysis.

The citric acid cycle: the pyruvate dehydrogenase complex: structure, coenzymes, regulation. Reactions of TCA cycle and regulation. The anaplerotic reactions.

Oxidative phosphorylation. Ultrastructure of mitochondria; redox potentials, respiratory chain complexes, chemiosmotic theory and electrochemical  potential. ATP synthase: structure and catalytic cycle. Inner membrane carriers, reducing equivalents shuttles. Energetic balance, inhibitors of oxidative phosphorylation, uncouplers. Reactive oxygen species and detoxifing enzymes

Gluconeogenesis: the pyruvate carboxylase; reactions of the pathway  in details, regulation of glycolysis and gluconeogenesis: role of PKA and PP1. Glycogen: structure, degradation and biosynthesis; insulin and glycogen metabolism.

Pentose phosphate shunt:
oxidative and non-oxidative phases: reactions and regulation. NADPH production and glutathione.

Lipid metabolism: utilization of triacylglycerols in animals, fatty acid activatio, transport and oxidation, keton bodies, fatty acid biosynthesis, metabolism of phospholipids containing glycerol, of sphingolipids, and of cholesterol.

Metabolism of aminoacids: protein turnover: chemical signals: ubiquitin, structure and activation, the proteasome. Utilization of ammonia: the urea cycle.

 

The experimental lab course on "Basic methods for separation, quantification and identification of proteins" (1CFU) will be held by Prof. Anna Maria Porcelli.  

It will cover:

Colorimetric determination of protein content;

Chromatographic separation of a mixture of known proteins;

Identification of purified proteins by means of spectrophotometric and elecrophoretic techniques.

Readings/Bibliography

Biochemistry, J. M. Berg, · J. L. Tymoczko, · L. Stryer , 7th edition 2012

Principes di Biochemistry. A.L. Lehninger, D.L. Nelson , M.M. Cox.

Foundamentals of Biochemistry:  Life at the Molecular Level, 2012, .D.Voet, J.Voet, C.W. Pratt. ,2013

PDF files of lectures slides will be available to students by downloading  from the AMS Campus web site upon registration to a distribution list and  use of a password.

Teaching methods

The lectures will focus on the structure and function of  major  molecules of biological interest and analize some important metabolic pathways and their integration.

Assessment methods

Written test comprising a series of questions that aim to ascertain students' understanding of the theory presented during lectures and experimental laboratory.

The written test comprizes:

Three open questions on general topics of the program; if not properly addressed, the evaluation will end (max 6 marks/corrected answer).

Four formulas of aminoacids, sugars, fatty acids, metabolites, etc. (1 mark/correct answer)

Five multiple-choice questions (1 mark/correct answer).

Three false-true questions (1 mark/correct answer)

Four problems similar to the ones dealt with during the practical exercises course (molarity, dilutions, Lambert-Beer law,  enzyme inhibition, etc (1 mark/correct answer).

 

Teaching tools

Ppt presentations

Office hours

See the website of Michela Rugolo

See the website of Anna Maria Porcelli