Enzyme Technology

Course Coordinator

ECTS points:
4

Program:
preddiplomski

Course number:
39780 (PDBT), 53709 (DBPI)

Course Description

COURSE CONTENT

1. Determination of enzyme kinetic parameters for the substrate conversion to product under conditions of enzyme industrial application

L: Thehistory of enzyme technology development. Sources of enzymes. Selection of enzymes for industrial application: determination of kinetic parameters and influence of certain parameters on enzyme reaction rate and process productivity (substrate concentration, enzyme concentration, Km as measure of enzyme affinity for substrate, enzyme turnover number, vmax). Influence of environmental parameters (pH, temperature, ionic strength) on enzyme activity and enzyme stability in conditions of enzyme industrial application. Applied kinetics of enzyme reactions. The influence of enzyme inhibitors and activators on the rate of enzyme reaction and their application in industrial processes.

 

2. Biotechnological production of free enzymes

L: Microbial biosynthesis of enzymes: medium, conditions, microorganisms, surface and submerged cultivation, surface cultivation methods. Enzyme isolation: general scheme, filtration, precipitation, extraction, concentration and cell disruption, chromatography, ultrafiltration and electrophoresis.

S: Biotechnological production of free enzymes for the industrial application

E: Wohlgemuth`s method. Anson`s method. Biosynthesis of a-amylase with Bacillus subtilis bacterium. Purification of active enzyme filtrate. SDS-PAGE electrophoresis of enzyme samples.

 

3. Biotechnological production of immobilized enzymes

L: Enzymes immobilization and stabilization. Methods of enzyme immobilization for industrial application. Application of membrane reactors in enzyme technology. Influence of environmental parameters and diffusion constraints on the immobilized enzyme kinetics.

S: Biotechnological production of free and immobilized enzymes for industrial application.

E: Immobilization of a-amylase in agar. Immobilization of alkaline proteases in calcium alginate. Determination of immobilized enzyme activity.

 

4. Industrial application of enzymes

L: Reactors and kinetic comparisons of enzyme reactors. A review of certain industrially important enzymes: proteolytic, amylolytic, pectinolytic, cellulose, penicillin-amidase and L-aminoacylase. Application of enzymes in biotechnological, food and other industries, analytical and scientific use, biosensors. Preparation and use of genetically modified microorganisms for the production of enzymes and enzyme-protein engineering. Implementation of thermophilic enzymes and enzymes in organic solvent. Legislation on the application of industrial enzymes in food production and as food additives.

S: Calculation of the required free/immobilized enzyme for implementation of the biocatalysis in industrial conditions. Application of enzymes in biotechnology, food and other industries.

LEARNING OUTCOMES

  • critically evaluate the advantages and disadvantages of using membrane bioreactors in enzyme technology
  • critically evaluate the enzyme for a particular industrial application based on the kinetic parameters of the enzyme reaction
  • critically evaluate the most suitable processes for the enzyme immobilization that will be used for substrate conversion in biotechnological products
  • critically evaluate the most suitable methods of isolation and purification of extracellularly and intracellularly produced enzymes
  • explain the influence of the environmental factors and diffusion constraints on the immobilized enzyme kinetics
  • explain the influence of environmental parameters (pH, temperature and ionic strength) and substrate concentrations on the enzyme activity and enzyme stability in conditions of enzyme industrial application
  • evaluate the success of immobilization methods by comparing the enzyme activity of proteolytic and amilolytic enzymes before and after immobilization
  • perform the biosynthesis, isolation and purification of enzymes by submerged cultivation of Bacillus subtilis bacteria
  • sketch the scheme of biotechnological process of enzyme production by submerged and surface cultivation of working microorganism
  • compare advantages and disadvantages of biotechnological production of enzymes by surface cultivation of microorganism on solid substrate with respect to submerged cultivation