Enzymology
Teacher
Dr Sitaram Swain
Category
Core Courses
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Course Name : Enzymology
Code(Credit) : CUTM1706(3-1-2)
Course Objectives
- • To acquire fundamental knowledge on enzymes and their importance in biological reactions.
• To understand ability to difference between a chemical catalyst and biocatalyst.
• Exposure to the nature of non-protein enzymes such as ribozymes.
• Understanding the role of enzymes in clinical diagnosis and industries.
Learning Outcomes
- • This course will provide fundamental knowledge on enzymes and their importance in
biological reactions.
• Students will understand the difference between a chemical catalyst and biocatalyst and understand activation energy.
• They will study non-protein enzymes such as ribozymes and will be exposed to the Industrial and biomedical applications of enzymes
Course Syllabus
Module:I Enzymes:Definition, historical perspective, general characteristics, co-factors – coenzymes and metal ions.
Module:II Classification and units of enzymes: Based on IUB with examples. Unit of enzyme activity,definition of IU, enzyme turn over number and nature of non-enzymatic and enzymatic catalysis. Specific activity. Enzyme specificity. Concept of active site, ES complex, specificity.
Module:III Theories of enzyme catalysis: Lock and key model, Koshland’s induced fit theory. Enzyme kinetics: Factors affecting rate of enzyme catalyzed reactions.
Module:IV Characterization: Effect of enzyme concentration, substrate concentration, pH and temperature. Michaelis – Menten equation,Lineweaver – Burk(L-B) plot. Determination of Vmax& Km from L-B plot and their significance.Enzyme inhibition –competitive, non competitive and uncompetitive. Graphical representation by L-B plot. Evaluation of Km, Ki and Vmax in presence of inhibitor.
Module:V Chemical modification of active site groups. Site directed mutagenesis of enzymes. Mechanism of action of chymotrypsin .
Module:VI Allosteric enzymes:Sigmoidal curve, positive and negative modulators, qualitative description of “concerted” & “sequential” models for allosteric enzymes. Half site reactivity, Flipflop mechanism, positive and negative co-operativity with special reference to aspartate transcarbamylase and phosphofructokinase.
Module:VII Isoenzymes:Detection, nature, importance. Lactate dehydrogenase as an example. Multi enzyme complex – Pyruvate dehydrogenase complex. – Composition, subunits, assembly, enzymatic reaction functions.RNA as an enzyme. (Ribozymes).
Applications of Enzymes: Enzymes as reagents, Marker enzymes in diagnostics, Immobilized enzymes, Industrial applications of enzymes.
Laboratory Practices
• Isolation of urease and demonstration of its activity
• Isolation of acid phosphatase and demonstration of its activity
• Determination of specific activity of salivary amylase by DNS
• Purification of urease
• Time course of urease reaction
• Influence of substrate concentration and pH on the rate of enzymatic reaction
• Determination of Km and Vmax of salivary amylase
• Determination of initial velocity [time kinetics] of salivary amylase
• Determination of optimum temperature of salivary amylase
Suggested readings:
Text Books:
Cox, M.M and Nelson, D.L. (2008). Lehninger Principles of Biochemistry, VEdition, W.H. Freeman and Co., New York.
Reference Books:
1.Cox, M.M and Nelson, D.L. (2008).Lehninger’s Principles of Biochemistry, Sixth Edition, W.H. Freeman and Co., New York.
2. Voet D and Voet J(2012) Biochemistry . Fifth edition, Wiley.
3. Murray, R.K., Bender, D.A., Botham, K.M., Kennelly, P.J., Rodwell, V.W. and Well, P.A. (2009). Harper’s Illustrated Biochemistry, XXVIII Edition, International Edition, The McGraw-Hill Companies Inc.
Session Plan
Session 1
Enzymes:Definition, historical perspective
Session 4
Classification and units of enzymes: Based on IUB with examples
Session 5
Unit of enzyme activity,definition of IU,
Session 6
enzyme turn over number and nature of non-enzymatic and enzymatic catalysis.
Session 7
Specific activity. Enzyme specificity
Session 8
Concept of active site
Session 9
ES complex
Session 10
specificity.
Session 11
Theories of enzyme catalysis: Lock and key model
Session 12
Koshland’s induced fit theory
Session 13
Enzyme kinetics: Factors affecting rate of enzyme catalyzed reactions
Session 14
Characterization: Effect of enzyme concentration, substrate concentration, pH and temperature.
Session 15
Characterization: Effect of enzyme concentration, substrate concentration, pH and temperature.
Session 16
Michaelis – Menten equation,Lineweaver – Burk(L-B) plot
Session 17
Determination of Vmax& Km from L-B plot and their significance.
Session 18
Enzyme inhibition –competitive, non competitive and uncompetitive.
Session 19
Graphical representation by L-B plot
Session 20
Evaluation of Km, Ki and Vmax in presence of inhibitor.
Session 21
Chemical modification of active site groups
Session 22
Site directed mutagenesis of enzymes
Session 23
Mechanism of action of chymotrypsin
Session 24
Allosteric enzymes:Sigmoidal curve, positive and negative modulators
Session 25
qualitative description of “concerted” & “sequential” models for allosteric enzymes.
Session 26
Half site reactivity
Session 27
Flipflop mechanism
Session 28
positive and negative co-operativity with special reference to aspartate transcarbamylase and phosphofructokinase
Session 29
Isoenzymes:Detection, nature, importance
Session 31
Multi enzyme complex – Pyruvate dehydrogenase complex.
Session 30
Lactate dehydrogenase as an example.
Session 31
Multi enzyme complex – Pyruvate dehydrogenase complex.
Session 31
Multi enzyme complex – Pyruvate dehydrogenase complex.
Session 31
Multi enzyme complex – Pyruvate dehydrogenase complex.
Case Studies
Case Studies
Our Main Teachers
Dr Sitaram Swain has received PhD degree from Utkal University in the year 2017. He is also trained and experienced from TATA Memorial Hospital, Mumbai, India. He has experienced with cell line culture, clinical biochemistry, immuno-fixation, immunoassay, cancer cytogenetic, serum protein electrophoresis and tumour marker analysis. He has qualified NET(ICAR-ASRB) in Animal biotechnology. He has […]