69162 - Chemical Structures Of Biological System

Learning outcomes

At the end of the courses the student will get the theoretical basis for understanding the relationship between the molecular structure of enzymes and the biochemical reactions catalyzed by them, particularly regarding the bioinorganic aspects of metal-enzymes and metallo-proteins. He will be able to acquire information, including those of spectroscopy nature, necessary to evaluate the chemical changes in the structure of molecules within the food

Course contents

Prerequisites

Students wishing to fully understand the course content should have a sound background in General and Inorganic Chemistry and in Organic Chemistry. In particular, they should be familiar with:

• the fundamentals of acid-base theory, including pH, pKa and chemical speciation;
• the Nernst equation and electrode potential;
• chemical kinetics, including the meaning of activation energy and the Arrhenius equation;
• molecular conformations and dihedral angles;
• nucleophilic and electrophilic attack.

Teaching Unit 1: Protein Structure (10 hours)

Descriptive elements of protein macromolecular structure (4 hours)

• Physicochemical properties of amino acids
• Primary structure of proteins
• Secondary structure and Ramachandran plot
• Prosthetic groups, apo- and holoproteins

Structural requirements for function (6 hours)

• Sequence homology: orthologues, paralogues and isoforms
• Metal ion-binding amino acids
• Metal coordination geometries and catalytic sites
• Dissociation and affinity constants
• Definition of catalytic site and mechanism of enzymatic reactions

Teaching Unit 2: Tools for the Acquisition and Processing of Structural Data (8 hours)

Structural elucidation of metalloproteins

• Instrumental methods: high-resolution NMR spectroscopy and X-ray crystallography for the acquisition of structural data
• NMR parameters and information on the chemical nature of biological systems
• Bioinformatics laboratory: software for the evaluation of protein structures, including MOLMOL and ClustalX

Teaching Unit 3: The Role of Metal Ions in Biological Systems (8 hours)

Bioavailability and distribution of metal ionic species in biological systems (4 hours)

• Abundance of metal ions in nature and in biological fluids
• Transport and bioavailability of metal ions
• Entry of metal ions into cells
• Transferrin, siderophores and chaperones
• Regulation of ion fluxes: enterobactin and competitive inhibition by ferric MECAM
• Lineweaver-Burk plot for the enterobactin-Fe³⁺ complex

Chemical kinetics and enzymology (4 hours)

• Parameters describing the rates of biological reactions
• Review of chemical kinetics
• Arrhenius equation
• Enzyme kinetics: Michaelis-Menten kinetics and saturation

Teaching Unit 4: Biological Functions of Metalloproteins (14 hours)

Hydrolytic enzymes (3 hours)

• Introduction to hard-soft acid-base theory
• Metal ions and the effect of coordination on the acid dissociation constant of water
• Enzymatic hydrolysis: role of metal ions in hydrolytic enzymes
• Mechanisms of action of carboxypeptidase and carbonic anhydrase

Chemical signal transduction (3 hours)

• Stabilising role of metal ions in macromolecular structure
• Ca²⁺ as a biological signal
• Affinity of protein ligands for calcium and magnesium ions
• Comparison between calmodulin and parvalbumin
• Coordination and conformational stabilisation
• Conformational flexibility and stability
• Transduction of chemical signals into mechanical energy: contractile proteins

Oxygen transport (4 hours)

• Structure-function relationships in oxygen transport
• Structural differences between haemoglobin and myoglobin and their implications for their distinct physiological functions
• Cooperativity and saturation curves of Hb and Mb, allostery and the Hill equation
• Myoglobin sequences in different species and affinity for O₂
• Spectroscopic properties of iron proteins coordinating other substrates
• Electronic states of iron ions in high-spin and low-spin configurations
• NMR of myoglobin and cyanide adducts
• Paramagnetic effects on the chemical shifts of NMR signals: the case of meat extracts
• Nuclear relaxation of water protons through interaction with paramagnetic ions: the case of egg-white ageing

Electron transport (4 hours)

• Thermodynamics of electron transfer
• Redox chemistry of water: reduction potentials of oxygen and H⁺
• Role of metal ions, Fe or Cu, and optimal coordination geometries in electron transport
• Role of ligand geometry in determining the E⁰ values of plastocyanin and azurin
• Kinetics of electron transfer
• Protein redox potentials and electron-transfer pathways
• Franck-Condon effect and Marcus theory
• Electron transport and catalysis
• Catalytic mechanism of superoxide dismutase
• Mobile Fe²⁺/Fe³⁺ ion in aconitase, maintenance of oxidative homeostasis, and analogies with IRE-BP

Readings/Bibliography

Scientific papers on authoritative journals (the list will be available on the Virtuale platform)

Bioinorganic Chemistry - I. Bertini, H.B. Gray, S.J. Lippard, J.S. Valentine (Eds.) -University Science Books, Mill Valley, CA, U.S.A., ISBN: 0935702571

Teaching methods

The course comprises 4 ECTS credits. Each ECTS credit includes 6 hours of lectures and 4 hours of practical activities carried out in the classroom or in the computer laboratory.

The computer laboratory is located at the Cesena Campus (Villa Almerici) and is equipped with 20 networked workstations. Students will be able to follow the procedures demonstrated by the lecturer via projection on a large screen.

The practical activities have two main objectives:

• to enable students to become familiar with software for the bioinformatic analysis of structural data;
• to develop students’ ability to formulate independently hypotheses concerning structure-function relationships in biological molecules.

Assessment methods

The final examination is designed to assess whether the following learning objectives have been achieved:

• the ability to use the information provided during the course to formulate hypotheses about the structural features of macromolecules;
• the ability to describe enzyme mechanisms using appropriate scientific language and rigour;
• the ability to define the criteria used to evaluate the effects of the chemical environment on the function of biological molecules.

Students may choose between a written examination and the independent preparation of a project accompanied by a PowerPoint presentation.

1. Written examination

The written examination consists of three open-ended questions on topics included in the course syllabus. Students will be asked to interpret and explain graphs, structural data and spectroscopic data provided during the examination.

Students who pass the written test will subsequently attend an oral discussion, during which they will be given the opportunity to explain and further discuss the content of their answers. The final grade will be determined upon completion of this oral discussion.

The assessment will take into account the appropriate use of scientific terminology, the accuracy and relevance of the answers, and the depth of the student’s knowledge. Concise answers are valued, as they help avoid unnecessary digressions.

2. Independently prepared project and slide presentation

Students may prepare an independent project on a topic of their choice among those included in the course syllabus. The project must be based on an original re-elaboration of data from the scientific literature and on structural information obtained using the software introduced during the course, namely MOLMOL and ClustalX.

The presentation should make effective use of the course content, whose slides are available on the Virtuale platform.

Any use of artificial intelligence must be critical and fully informed. All figures, graphs and information included in the presentation must be checked for accuracy and fully understood by the student. The inclusion of unverified material or material that the student is unable to explain adequately will be regarded negatively in the assessment.

During the examination, students must therefore be able to explain not only what is shown in the slides, but also the reasoning underlying the presentation, the bibliographic sources used and their own personal contribution.

The slide-based examination is intended to provide an in-depth exploration of the topics covered during the course. Slides should not contain excessive amounts of text, but should instead focus on key concepts, diagrams and images that support the discussion.

Where appropriate to the selected topic, the presentation should include:

• a ClustalX comparison of at least ten homologous sequences, including, where possible, orthologues from different species and paralogues belonging to the same protein family, such as different enzyme isoforms, together with a discussion of the functional significance of conserved residues and observed sequence variations;
• the localisation, on a three-dimensional structure prepared using MOLMOL, of the key amino acids involved in protein function;
• conformational analysis of the three-dimensional structure through φ–ψ rotations, with discussion of their structural implications;
• an explanation of the protein’s molecular mechanism, highlighting the role of the amino acids directly responsible for its biological function;
• minimal use of previously published figures, giving preference instead to analyses and representations prepared independently by the student, so as to demonstrate their bioinformatics skills;
• a discussion of the role of any metal ions present in the structure. Metalloproteins often provide particularly informative case studies and are therefore strongly recommended.

The information used should be drawn primarily from the scientific literature, including original research articles and review papers, rather than from general-interest websites.

The lecturer is fully available to provide any clarification required, including through more than one online meeting, before the examination and presentation date is agreed.

Students with specific learning disorders, temporary disabilities or permanent disabilities are advised to contact the relevant University office well in advance. The office will propose any appropriate adjustments, which must in all cases be submitted to the lecturer for approval at least 15 days before the examination. The lecturer will assess their suitability in relation to the learning objectives of the course.

Teaching tools

The teaching materials presented during lectures are made available to students in electronic format.

The free software packages ClustalX and MOLMOL can be downloaded from the Virtuale platform.

Audio recordings of lectures are permitted, provided that they are made available to all students by sharing the MP3 files in restricted-access areas of the lecturer’s website.

The course also includes access to public protein databases.

Office hours

See the website of Francesco Capozzi