Protein Purification and Characterisation

Course Coordinator

ECTS points:
4

Program:
preddiplomski

Course number:
39806

Course Description

COURSE CONTENT

Lectures: Isolation and solubilisation of proteins from natural materials. Separation of proteins by precipitation techniques. Separation of proteins by centrifugation and by using semipermeable membranes. Separation of proteins by chromatographic methods (gel filtration, ion exchange, biospecific (affinity) chromatography). Genetic tagging of proteins for purification. Electrophoretic methods (polyacrylamide gel electrophoresis, immunoblot, isoelectric focusing). Application of analytical methods for protein characterization (NMR, MS, CD spectroscopy, X-ray crystallography). Designing of protein purification procedures.

Seminars: Preparation for practical courses. Computer simulation of the protein purification process.

Practical courses: Purification of ovalbumin from hen egg to homogeneity by ammonium sulphate precipitation, ion exchange chromatography and gel filtration. Testing the sample for purity by electrophoresis after each purification step. Purification of yeast acid phosphatase to homogeneity by ammonium sulphate precipitation, ion exchange chromatography and gel filtration. Analysis of specific activity after each purification step.

LEARNING OUTCOMES

  • propose the protein purification procedure starting from natural materials or cultivated biomass
  • select and conduct common preparative methods for protein separation; identify advantages, limitations and capabilities of each method
  • select and conduct common analytical methods for protein separation in order to monitor the progress of purification and apply appropriate analytical methods for characterization of purified proteins
  • evaluate efficiency of the purification procedure and modify it in order to increase efficiency of the method, yield or quality of the final product and decrease the cost of the purification procedure

To enrol in this course, the following courses must be completed:

  • Biochemistry 1