00061 - Biochemistry

Learning outcomes

At the end of the course, the student will have knowledge of: biological processes at the molecular level; structure-function relationships of biomolecules, with particular focus on proteins; energy metabolism; an integrated view of signaling pathways and major metabolic processes; the fundamentals of structural biochemistry and enzymology; the main spectrophotometric methods for protein quantification; electrophoretic separation techniques and protein identification methods; the use of experimental protocols for calculating enzyme specific activity. Upon completion of the laboratory component, the student will also be able to critically evaluate the results obtained by preparing a report on the experiments performed.

Course contents

Module 1 (6 ECTS) Biochemistry (Instructor: Concettina Cappadone)

 PROTEIN STRUCTURE–FUNCTION RELATIONSHIPS Hemoglobin and myoglobin. Structure of the heme group. Oxygen saturation curves and cooperativity. Allosteric effectors.

FUNDAMENTALS OF BIOENERGETICS Thermodynamics and spontaneity of metabolic processes. Definition and biological significance of free energy; relationships between free energy, entropy, enthalpy, and the equilibrium constant. Mechanisms of energy transfer: coupled reactions. High-energy biological molecules: high-energy phosphorylated compounds and reduced nucleotides. ATP and the phosphoryl group transfer potential.

ENZYMOLOGY Enzymes: classification. Enzyme catalysis. Activation energy. Structure and general properties of enzymes. Cofactors and coenzymes. Enzyme kinetics: Km, Vmax, kcat. The Michaelis–Menten equation. Effects of pH and temperature on enzyme activity. Enzyme inhibition: reversible competitive and non-competitive inhibition. Regulation of enzyme activity: allosteric enzymes and cooperativity, positive and negative modulators; reversible, irreversible, and covalent modifications.

METABOLISM General organization and compartmentalization of metabolic pathways. Catabolism and anabolism: relationship between degradation and biosynthetic pathways; substrate cycles. Definition of rate-limiting steps in metabolic pathways, control mechanisms, and the role of ATP in determining pathway spontaneity. Unidirectionality of metabolic pathways and regulation of metabolic flux.

CARBOHYDRATE METABOLISM Glycolysis: fate of pyruvate under aerobic and anaerobic conditions; glycolytic reactions and their regulation. Pentose phosphate pathway. Gluconeogenesis and glycogen metabolism. Coordinated regulation of glycogen synthesis and breakdown. Gluconeogenesis: substrates and reactions; regulation mechanisms. Overview of the metabolism of other monosaccharides (fructose, galactose).

CITRIC ACID CYCLE AND OXIDATIVE PHOSPHORYLATION Production of acetyl-Coenzyme A. Pyruvate dehydrogenase complex and its five cofactors. Regulation mechanisms of pyruvate dehydrogenase. Citric acid cycle: reactions, regulatory mechanisms, and metabolic interconnections (role of anaplerotic reactions). Electron transport and oxidative phosphorylation: 3D structure of electron carriers in the mitochondrial respiratory chain. ATP synthesis: role of the transmembrane proton gradient. Structure of ATP synthase. Regulation of oxidative phosphorylation: cellular energy demand and the role of uncouplers in thermogenesis. Hypoxia and generation of reactive oxygen species (ROS).

LIPID METABOLISM  Digestion, mobilization, and transport of fatty acids. Fatty acid catabolism: β-oxidation reactions. Formation and utilization of ketone bodies. Fatty acid biosynthesis: reactions and regulation. Fatty acid elongation and desaturation. Overview of triglyceride and phospholipid biosynthesis. Cholesterol, steroids, and isoprenoids: biosynthesis, regulation, and transport.

AMINO ACID DEGRADATION AND UREA PRODUCTION Metabolic fate of amino groups: transamination and oxidative deamination reactions. Urea cycle: localization of reactions and interconnections between the urea cycle and the citric acid cycle.
INTRODUCTION TO SIGNAL TRANSDUCTION  cAMP and phosphoinositide-dependent signaling pathways. Tyrosine kinase receptors and phosphorylation cascade mechanisms.

Module 2 (2 ECTS) – BIOCHEMISTRY LABORATORY

The 2 ECTS credits of the Biochemistry Laboratory module represent an integrated practical component within the Biochemistry course. The laboratory is designed to introduce students to the fundamentals of enzyme study. Students will learn how to calculate the specific activity of an enzyme in a cellular protein lysate. They will evaluate the kinetic parameters of a purified enzyme, examine the pH dependence of enzyme activity, and assess substrate specificity.

Specifically, the laboratory includes the following experiments:

1. Determination of protein content in a protein extract using the Bradford assay and calculation of the specific activity of an enzyme.

2. Determination of the kinetic parameters Km and Vmax of the enzyme alkaline phosphatase.

3. Investigation of the effect of pH and substrate specificity on trypsin activity.

4. Determination of the molar extinction coefficient of NADPH using the glucose-6-phosphate dehydrogenase reaction.

5. Analysis of cellular lipids through staining of slides with cells of different histogenesis.

At the end of the laboratory, a data analysis session will be held, during which students will evaluate the results both individually and in groups. The principles of writing a scientific report will be discussed based on the experimental outcomes.

In addition to the practical activities, brief theoretical lectures will be provided on methods for protein extraction from biological samples, electrophoresis, and UV-Vis absorption spectroscopy.

Readings/Bibliography

Berg, Tymoczko, Stryer "Biochimica" Zanichelli editore

Nelson, Cox "I principi di Biochimica di Lehninger" Zanichelli editore

Garret e Grisham "Biochimica" V edizione Piccinin editore

Biochimica Molecole e metabolismo • Con Mastering Chemistry™ Dean R. Appling - Spencer J. Anthony-Cahill - Christopher K. Mathews Pearson editore

Laboratory Materials:

Students will be provided with handouts detailing the procedures for each individual experiment. All materials used during the theoretical lessons will also be made available. No specific textbook is required.

Teaching methods

Module 1: Lectures 
Module 2: Individual laboratory work and group data analysis. Attendance in the laboratory is mandatory.

"Given the nature of the activities and the teaching methods adopted, participation in this learning activity requires all students enrolled in Modules 1 and 2 to complete the e-learning course on safety and health in study and training environments [https://www.unibo.it/en/services-and-opportunities/health-and-assistance/health-and-safety/health-and-safety-in-study-and-training-places ]. Additionally, students must attend Module 3, a specific training course on safety and health in study environments. Information regarding the dates and attendance procedures for Module 3 is available in the dedicated section of the degree program website."

Assessment methods

The final assessment aims to evaluate the achievement of the following learning objectives:

Module 1 – Biochemistry

-Understanding the structure and function of the main biological macromolecules and the fundamental principles of enzymology.

-Understanding cellular bioenergetics, major metabolic pathways, and the underlying mechanisms of their regulation and signal transduction.

-Understanding the basic molecular mechanisms of living systems and the molecular logic behind their regulation.

Module 2 – Biochemistry Laboratory

-Knowledge of the main methods for protein quantification.

-Knowledge of experimental methods for evaluating enzyme activity.

The examination includes:

-A written report on the laboratory activities of Module 2.

-An oral examination to assess Module 1 (Biochemistry).

The oral exam consists of three questions aimed at verifying the student’s theoretical understanding of the structure and function of biological macromolecules, metabolism, and its regulation.

Throughout the laboratory activities, students will be guided by the instructors of both modules and supported by tutors. Continuous interaction will be encouraged to clarify the analytical approach and individual experiments.

To pass the laboratory module, students must submit a written report based on the format presented during the final lab session. The report must be delivered within one week after the end of the laboratory.

The final grade, expressed on a 30-point scale, will be the weighted average of the grades obtained in the two modules.

Teaching tools

Module 1: PowerPoint presentations.

Module 2: Practical laboratory equipped with spectrophotometers, thermostatic water baths, electrophoresis cells, and optical microscopes. Computer lab with individual workstations.

 

Students with learning disorders and\or temporary or permanent disabilities: please, contact the office responsible (https://site.unibo.it/studenti-con-disabilita-e-dsa/en/for-students ) as soon as possible so that they can propose acceptable adjustments. The request for adaptation must be submitted in advance (15 days before the exam date) to the lecturer, who will assess the appropriateness of the adjustments, taking into account the teaching objectives.

Office hours

See the website of Concettina Cappadone

See the website of

See the website of