- Assessment methods
- Learning objectives
- Teaching methods
The student who attends this course will be asked to apply the knowledge acquired in several subjects: Biochemistry, in particular solid knowledge regarding the structure and properties of proteins are required; Molecular Biology, with particular reference to recombinant DNA techniques (manipulation, amplification, cloning) but also to gene expression regulation mechanisms; Microbiology, as regards the distinctive features of the microorganisms used as heterologous hosts for the production of recombinant proteins and their maintenance in culture. In order for the student to be able to carry out the experimental work during the planned laboratory activities, the knowledge and practical skills acquired during the course of Biochemical Methodologies are required.
The exam is organized in a written test concerning the modules of “Protein Engineering” and “Laboratory of Recombinant Proteins” (four open questions for each module). Laboratory activities will be evaluated through a written scientific report of the experimental procedures and results. The final score is given in 30ies: the examination is successful if a score 18/30 is gained in each module and in the laboratory activities report.
This course of Protein Engineering and Laboratory Recombinant Proteins is fundamental to the preparation of a student in Industrial and Molecular Biotechnology. The aim of the course is to explain the molecular mechanisms at the basis of the structure-function relationships of proteins and the experimental approaches to modulate the protein functionality and to evolve a desired function or structure. The course is also aimed to discuss the most updated knowledge/skills related to the production of recombinant proteins. It therefore provides the theoretical tools for the design and production in different heterologous systems of protein variants.
In particular, the Protein Engineering module aims to provide the knowledge of the different techniques in the biotechnological application of protein engineering: design of protein structures from scratch, rational design for the design of new functions, molecular modeling and docking, mutagenesis (site-directed and random) as well as methods through which it is possible to isolate the optimized protein variants in a given property.
The Laboratory of Recombinant Proteins module is designed to provide an overview of the different expression systems used in the production of recombinant proteins for different applications (industrial or pharmaceutical). For each system the most recent methods for the preparation of expression constructs, the regulation of protein expression levels and the optimization of production yields will be exposed.
Students will be encouraged to evaluate the advantages and limitations of the various approaches used to modify proteins properties and those related to the use of different systems of heterologous expression. Case studies will be also discussed. The course includes laboratory activities during which the theoretical contents will be applied experimentally. Students are expected to acquire decision-making skills in a "protein engineering" project, from the design phase to the production of recombinant protein variants one, as well as to be able to face and solve the various problems related to the process.
Learning outcomes can be summarized as follows.
- acquisition of fundamental elements at molecular level concerning protein structure-function relationships;
- acquired the required information (theoretical and experimental) to carry out the process of engineering of a protein function/structure.
- ability to read, understand and criticize a scientific text on the production, characterization and engineering of recombinant proteins (in English);
- ability to use these knowledge to evaluate the aims and/or the results of a research project both on the qualitative and quantitative point of view.
Practical and subject specific skills:
- ability to apply the knowledge in the field of biochemistry and molecular biology to a protein engineering project;
- ability to propose an analytical problem-solving approach.
- ability to identify and express relevant information;
- demonstrate effective communication skills by practicing, reading, writing and speaking clearly;
- demonstrate the ability to resume and present the scientific information.
Module “Protein Engineering” – Loredano Pollegioni: 6 CFU – 48 hours class
Structure-function relationships in proteins (4 CFU, 32 hours)
- Introduction to protein structure
- Protein classification
- Protein synthesis
- Methods for protein structure determination
- Post-translational modification and protein targeting
- Protein folding
- Protein degradation
- Membrane proteins
Protein engineering (2 CFU, 16 hours)
- De novo protein design
- Rational design (with examples)
- Molecular docking
- Directed evolution (with examples)
- Screening methods
- Engineering of protein stability
Slides of the lessons: on the e-learning website
Arnold F. H., Georgiou G. Directed Enzyme Evolution: Screening and Selection Methods (Methods in Molecular Biology) (Humana Press)
Arnold F. H., Georgiou G. Directed Evolution Library Creation: Methods and Protocols (Methods in Molecular Biology) (Humana Press)
Branden C. e Tooze J., Introduction to protein structure (Garland Publ.)
Bross P. e Gregersen N. Protein misfolding and disease, Methods in Molecular Biology, vol. 232 (Humana Press)
Garette-Grisham "Principi di Biochimica" (PICCIN)
Matthews, Van Holde, Ahern, “Biochimica” (Casa Editrice Ambrosiana)
Voet D, Voet JG, Pratt CW "Fondamenti di Biochimica" (Zanichelli)
Module “Protein Engineering” (L. Pollegioni) 6 CFU - 48 h class
Students can meet with the professor in his office by previous email appointment.