Enrollment year
2017/2018
Academic discipline
BIO/10 (BIOCHEMISTRY)
Department
DEPARTMENT OF BIOLOGY AND BIOTECHNOLOGY "LAZZARO SPALLANZANI"
Period
2nd semester (01/03/2020 - 14/06/2020)
Lesson hours
60 lesson hours
Activity type
WRITTEN AND ORAL TEST
Prerequisites
For a clear understanding of the structural and functional properties of proteins and enzymes, the knowledge of the principal bases of biochemistry is required.
Learning outcomes
The main aim of the course is to provide the basical knowledges about structure, mechanisms of action, regulation and biotechnological applications of the enzymes.
Course contents
General Enzymology (3 CFU)
Structure and function relationship of the enzymes. Lock and key model, and induced fit model. The active site study: covalent labeling and affinity labeling; double labeling; quasi-substrate inhibitors, suicide inhibitors. Catalysis strategies: proximity and orientation, induced fit, covalent nucleophilic catalysis, general and specific acid-base catalysis. Structure and mechanism of action of lysozyme, glutathione reductase, chymotrypsine. Enzyme kinetics: determination of the initial velocity (Vo), steady-state theory, Michaelis-Menten equation, Lineweaver-Burk method for Vmax and Km determination. Enzyme inhibition: competitive and non competitive inhibition. Enzyme activity regulation: effects of pH and temperature; allosteric enzymes, sequential transition and concerted transition models, homotropic and heterotropic effects. Aspartate transcarbamilase. Covalent regulation of enzymes. The serine proteases of the coagulation cascade: thrombin, structure, mechanism of action, mechanisms of activation and inhibition. Isoenzymes.
Laboratory of Applied Enzymology (3 CFU)
Method of extraction, purification and characterization of the enzymes. The laboratory experience consists in the purification of an enzyme, and in the characterization of its main kinetic properties. The main technique approached are: preparation of buffers and solution; chromatographic techniques for protein purification, centrifugation, electrophoresis, protein and enzymatic assays, biochemical calculations for the determination of the kinetic parameters. The aim of this module is to provide the main knowledge and skill for enzyme investigation, and to understand the potential of enzyme catalysis in industrial and biomedical applications. The different biochemical techniques will be approached both theoretically and practically.
Teaching methods
Module General Enzymology: frontal lessons.
Module Laboratory of Applied Enzymology: a series of experiments conducted in educational laboratories, accompanied by short theoretical lessons on applied techniques.
Reccomended or required readings
I aGeneral Enzymology (3 CFU)
Lesson notes.
Texts: the same ones used for Biochemistry nd II courses (chapters about the biochemistry of enzymes).
Laboratory of Applied Enzymology (3 CFUs)
Lesson notes and provided material.
Reference texts for possible integrations: I principi di Biochimica di Lehninger (Nelson e Cox), Fondamenti di Biochimica (Voet, Voet, Pratt)
Assessment methods
Oral exam which includes the discussion of the written report about the laboratory experience of the Laboratory of Applied Enzymology Module
Further information
Students are required to bring a lab coat to wear during the experiments.
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