Biochemistry and analytical biochemistry (2016/2017)

Course code
4S02696
Credits
12
Coordinator
Paola Dominici
Other available courses
Teaching is organised as follows:
Unit Credits Academic sector Period Academic staff
BIOCHIMICA E BIOCHIMICA ANALITICA: BIOCHIMICA 8 BIO/10-BIOCHEMISTRY I sem. Paola Dominici
BIOCHIMICA E BIOCHIMICA ANALITICA: BIOCHIMICA ANALITICA 4 BIO/10-BIOCHEMISTRY See the unit page See the unit page

Learning outcomes

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MM: BIOCHIMICA E BIOCHIMICA ANALITICA: BIOCHIMICA
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By the end of the course students should be able to: 1. Understand the basis for the molecular structure of different biochemical compounds; 2. Understand the biosynthesis of basic biochemical “building blocks”; 3. Understand the conformation, dynamics, and function of proteins; 4. Understand the generation and storage of metabolic energy; 5. Understand overall aspects of the integration of metabolic processes; 6. Have developed the skills to accumulate, integrate, and apply biochemical information in their own field of study
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MM: BIOCHIMICA E BIOCHIMICA ANALITICA: BIOCHIMICA ANALITICA
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The course is designed to provide the basic knowledge of the main biochemical methods used in the separation, identification, characterization and analysis of biomolecules, with particular reference to applied strategies used for the study of proteins.

Syllabus

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Course Description: Chemistry of biomolecules with emphasis on the bioenergetics and control of metabolic pathways via regulation of specific enzymes. Specific Coverage: I. Proteins, protein structure and mechanism. II. Enzyme mechanism and Enzyme kinetics III. Metabolism: Carbohydrates, conversion of sugars, anaerobic and aerobic metabolism. Fat catabolism. Amino acid degradation. IV. Anabolic Pathways: Carbohydrates, fats and lipids, amino acids and steroids. V. DNA replication
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Protein purification pH-buffer solutions: choice of buffers for biochemistry. Amino acids and isoelectric point. Homogenization: choice of methods and media. Fractional precipitation of proteins: fractional precipitation with ammonium sulfate and isoelectric precipitation with organic solvents and polymers; heat precipitation. Dialysis and ultrafiltration: principles and applications. Purification of enzymes: specific activity and yield. Spectroscopic techniques UV-VIS in Biochemistry. Report of Lambert-Beer. Colorimetric assays. Spectrofluorimetry (general principles and applications) and Chemiluminescence. Fluorescent proteins. Fluorescence resonance energy transfer (FRET). Centrifugal techniques Principles of sedimentation, RCF, sedimentation coefficient and sedimentation times. Centrifuges and rotors. Differential centrifugation: separation and analysis of subcellular fractions. Density gradient centrifugation, zonal and isopycnic. Applications: separation of cells, subcellular organelles, proteins, nucleic acids. Chromatographic techniques General principles, resolution and theoretical plates. Column chromatography. Hydrophobic chromatography of proteins (HIC).Ion-exchange chromatography of proteins. Exclusion chromatography (gel filtration). Affinity chromatography. Immuno-affinity chromatography. Chromatography with dyes and lectins. Chromatography with metals; purification of histidine-tagged proteins and of recombinant proteins. Electrophoretic techniques General principles, factors that influence the electrophoretic mobility. Gel Electrophoresis: PAGE, SDS-PAGE, Agarose. Methods of detection and quantitative assessments. Blotting -Western blotting in the study of proteins. Isoelectric focusing (IEF). Capillary electrophoresis. Two-dimensional electrophoresis. Practical laboratory excercises 1. Determination of protein concentration by absorbance at 280 nm a nd by colorimetric assay. 2. Determination of the NADH absorbance UV-Vis spectrum and determination of the extinction coefficient of NADPH 3. Determination of kinetic parameters (vmax and Km) of acid phosphatase 4. Determination of protein molecular weight by size-exclusion chromatography 5. Gel Electrophoresis SDS-PAGE 6. Western blotting 7. Oral presentation of a scientific journal paper

Assessment methods and criteria

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MM: BIOCHIMICA E BIOCHIMICA ANALITICA: BIOCHIMICA
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Modalita’ esame: L’esame finale (scritto per gli studenti di Bioinformatica) verterà su tutti gli argomenti del programma. Lo studente dovrà rispondere ad una serie di domande a risposta aperta, dimostrando di aver compreso e di essere in grado di utilizzare i concetti fondamentali di ogni argomento. L’esame finale (orale per gli studenti di Biotecnologie) verterà su tutti gli argomenti del programma. Lo studente dovrà rispondere ad una serie di domande, dimostrando di aver compreso e di essere in grado di utilizzare i concetti fondamentali di ogni argomento.
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MM: BIOCHIMICA E BIOCHIMICA ANALITICA: BIOCHIMICA ANALITICA
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The exam consists of a written examination and a facultative oral examination