M.Sc.Microbiology Semester III-

TH-IX ENZYME TECHNOLOGY

Unit – 1 Extraction and purification of microbial enzymes
Importance of enzyme purification, different sources of enzymes. Extracellular and intracellular enzymes. Physical and Chemical methods used for cell disintegration. Enzyme fractionation by precipitation (using Temperature, salt, solvent, pH, etc.), liquid-liquid extraction, ionic exchange, gel chromatography, affinity chromatography and other special purification methods. Enzyme crystallization techniques. Criteria of purity of enzymes. Pitfalls in working with pure enzymes.

Unit – 2 Enzyme inhibition and Co-factors
Irreversible, reversible, competitive, non-competitive and un-competitive inhibition with suitable examples and their kinetic studies.
Allosteric inhibition, types of allosteric inhibition and their significance in metabolic regulation & their kinetic study Vitamins and their co-enzymes: structure and functions with suitable examples. Metalloenzymes and Metal ions as co-factors and enzyme activators.

Unit – 3 Immobilization of microbial enzymes
Methods viz. adsorption, covalent bonding, entrapment & membrane confinement and their analytical, therapeutic & industrial applications. Properties of immobilized enzymes.

Unit – 4 Enzyme Engineering
Chemical modification and site-directed mutagenesis to study the structure-function relationship of industrially important enzymes.

Unit – 5 Applications of microbial enzymes
Microbial enzymes in textile, leather, wood industries and detergents. Enzymes in clinical diagnostics.
Enzyme sensors for clinical processes and environmental analyses. Enzymes as therapeutic agents.

References-

  1. Methods in Enzymology. Volume 22 – Enzyme purification and related techniques. Edited by William B. Jakoby. Academic Press, New York.
  2. Allosteric Enzymes – Kinetic Behaviour. 1982. by B.I. Kurganov. John Wiley and Sons. Inc., New York.
  3. Biotechnology. Volume 7 A – Enzymes in Biotechnology. 1983 Edited by H. J. Rehm and G. Reed. Verlag Chemie.
  4. Hand Book of Enzyme Biotechnology by Wiseman.
  5. Enzymes as Drugs Edited by John S.Holcenberg and Joseph Roberts , John Wiley & Sons New York.
  6. Methods of Enzymatic Analysis by Hans Ulrich, Bergmeyer, Academic Press.
  7. Methods in Enzymology by W.A. Wood, Academic Press.
  8. Advances in Enzymology by Alton Meister, Interscience Publishers.
  9. Topics in Enzyme and Fermentation Biotechnology by L.N. Wiseman, John Wiley and Sons.

TH X BIOPROCESS ENGINEERING AND TECHNOLOGY

Unit–1 Bioreactors
Design of a basic fermenter, bioreactor configuration, design features, individual parts, baffles, impellers, foam separators, sparger, culture vessel, cooling and heating devices, probes for online monitoring, computer control of fermentation process, measurement and control of process. Reactors for specialized applications: Tube reactors, packed bed reactors, fluidized bed reactors, cyclone reactors, trickle flow reactors, their basic construction and types for distribution of gases.

Unit – 2 Mass transfer in reactors
Transport phenomena in fermentation: Gas- liquid exchange and mass transfer, oxygen transfer, critical oxygen concentration, determination of Kla, heat transfer, aeration/agitation, its importance. Sterilization of Bioreactors, nutrients, air supply, products and effluents, process variables and
control, scale-up of bioreactors.

Unit – 3 Fermentation process
Growth of cultures in the fermenter Importance of media in fermentation, media formulation and modification.
Kinetics of growth in batch culture, continuous culture with respect to substrate utilization, specific growth rate, steady state in a chemostat, fed-batch fermentation, yield of biomass, product, calculation for productivity, substrate utilization kinetics.
Fermentation process: Inoculum development. Storage of culturesfor repeated fermentations, scaling up of process form shake flask to industrial fermentation.

Unit – 4 Down stream processing
Biomass separation by centrifugation, filtration, flocculation and other recent developments.
Cell disintegration: Physical, chemical and enzymatic methods.
Extraction: Solvent, two phase, liquid extraction, whole broth, aqueous multiphase extraction. Purification by different methods.
Concentration by precipitation, ultra-filtration, reverse osmosis. Drying and crystallization.

Unit – 5 Microbial strain improvement
Isolation, selection and improvement of microbial cultures: Screening and isolation of microorganisms, primary and secondary metabolites, enrichment, specific screening for the desired product.
Strain improvement for the selected organism: mutation and screening of improved cultures, random and strategic screening methods, strategies of strain improvement for primary, secondary metabolites with relevant examples. Use of recombinant DNA technology, protoplast fusion techniques for strain improvement of primary and secondary metabolites. Production of recombinant molecules in heterologus system, problems associated with strain improvement programme, improvement of characters other than products and its application in the industry.
Preservation of cultures after strain improvement programme.

References-

  1. Principles of Fermentation Technology by Stanbury, P.F., Whitekar A. and Hall. 1995., Pergaman. McNeul and Harvey.
  2. Fermentations – A practical approach. IRL.
  3. Bioprocess Technology: Fundamentals and Applications. Stockholm KTH.
  4. Biochemical Reactors by Atkinson B., Pion, Ltd. London.
  5. Biotechnology – A Text Book of Industrial Microbiology by Cruger.
  6. Fermentation Biotechnology: Industrial Perspectives by Chand.
  7. Biochemical Engineering Fundamentals by Bailey and Ollis, Tata McGraw Hill, N.Y.
  8. Biotechnology. Volume 3. Edited by H. J. Rehm and G. Reed. Verlag Chemie. 1983.
  9. Advances in Biochemical Engineering by T.K. Bhosh, A.Fiechter and N. Blakebrough. Springer Verlag Publications, New York.
  10. Biotechnology- A textbook of Industrial Microbiology by Creuger and Creuger, Sinaeur Associates.
  11. Bioprocess Engineering Kinetics, Mass Transport, Reactors, and Gene expressions by Veith, W.F., John Wiley and Sons.
  12. Applied Microbiology Series.
  13. Industrial Microbiology by L.E. Casida, Wiley Eastern
  14. Bioseparation: Down stream processing for Biotechnology by Belter, P.A. Cussler, E.L. and Hu, W.S., John Wiley and Sons, N.Y.
  15. Separation process in Biotechnolgy by Asenjo, J.A. Eds. Marcel Dekkar, N.Y.
  16. Bioprocess Engineering Principles by Doran, Acad. Press, London.
  17. Bioreaction Engineering Principles by Nielsen, J. and Villadsen, plenum Press, N.Y.
  18. Fermentation, Biocatalysis and bioseparation, Encyclopedia of Bioprocess Technology by Chisti, Y., Vol. 5, John Wiley and Sons, N, Y.

TH-XI MICROBIAL GENETICS

Unit – 1 DNA Structure and Mutagenesis
Historical developments in genetics, discovery of DNA and experimental evidence, Structure of Circular DNA molecule, Primary, Secondary, Tertiary and Quaternary structure of DNA, Watson and Crick model of double stranded DNA the law of DNA constancy and C value paradox and
topological manipulations.
DNA replication: DNA replication mechanism, enzymes involved in DNA replication and models of DNA replication. Molecular basis of spontaneous and induced mutations [physical and chemical mutagenic agents], types of mutation: point, frameshift, lethal, conditional lethal, inversion and deletion, null mutation, reversion of mutations, intra and intergenic suppression mutations. Environmental mutagenesis,
toxicity testing and population genetics. Systems that safeguard DNA. DNA methylation and DNA repair mechanisms – excision, mis- match, SOS , photoreactivation, recombination repair and glycocylase system.

Unit – 2 Prokaryotic Transcription and Translation
Organization of transcriptional units and regulation of gene expression Mechanism of transcription of prokaryotes-Structure and function of RNA polymerase, [DNA foot printing], termination and antitermination – N proteins and nut sites in DNA binding proteins, enhancer sequences and control of transcription, RNA processing (Capping, polyadenylation, splicing, introns and exons) Ribonucleoprotein, structure of mRNA, rRNA, tRNA. Direction of protein synthesis, RNA template, direction with experimental proof, tRNA as adaptor, ribosomes and their organization in prokaryotes, polycistronic mRNA in bacteria, initiation of translation in bacteria, small sub-units, its accessory factors, SD sequence in bacteria, initiator tRNA, elongation of translation, translocation and termination mechanisms. Post-translationalmodification. Salient features of genetic code.

Unit – 3 Regulation of gene expression in prokaryotes
Operon concept, co-ordinated control of structural genes, stringent response, catabolite repression, instability of bacterial RNA, positive regulation in E.coli [Arabinose operon] and negative regulation in E.coli [lac operon], inducers and repressors, regulation by attenuation bytrp operon.

Unit – 4 Genetic recombination
Genetic recombination processes: Role of rec proteins in homologous recombination.
Conjugation: Discovery, F+, F– and Hfr cells, types of Hfr; F+ and F– and Hfr and F– genetic crosses. Mechanism of conjugation. Sexduction, conjugational transfer of colicinogenic and resistance transfer factors. Genetic mapping. Plasmid Replication and Incompitability, Control of
copy number. Transposons – Insertion sequences and composite transposons, phages as transposons, replicative, non-replicative and conservative transposition. Mutations i.e. deletions, inversions and frame- shift due to transposition. Mechanism of transposition, controlling elements of maize – autonomous and non-autonomous elements. Types of transposons and their properties.

Unit – 5 Phage Genetics
T4 virulent phage: structure, life cycle, genetic map and DNA replication. Lamda temperate phage: Structure, genetic map, lytic and lysogenic cycle, lysogenic repression and phage immunity. [Lambda regulon] applications of phages in microbial genetics.

References-

  1. Microbial Genetics by Maloy ET. Al. 1994. Jones and Bartlett Publishers.
  2. Molecular Genetics of Bacteria by J. W. Dale. 1994. John Wiley and Sons.
  3. Modern Microbial Genetics. 1991 by Streips and Yasbin. Niley Ltd.
  4. Moleculat Biology of the Gene 4th Edition by J.D. Watson, N.H. Hoppkins, J.W. Roberts, J.A. Steitz and A.M. Weiner. 1987, The Benjamin / Cummings Publications Co. Inc. California.
  5. Gene VII by Lewin Oxford University Press. 2000.
  6. Bacterial and Bacteriophage Genetics. 4th Editions by Birge.
  7. Microbial Genetics by Frefielder. 4th Edition.
  8. Organization of Prokayotic Genome. 1999 by Robert L.Charlebois, ASM Publications.
  9. DNA repair and mutagenesis. 1995 by Errol C. Friedberg, Graham C. Walker and Wolfram, Siede, ASM Publications.
  10. Molecular Genetics of Bacteria, 1997 by Larry, Snyder and Wendy, Champness, ASM Publications.
  11. Methods of General and Molecular Bacteriology, 1993. Edited by Philip. Gerhardt, ASM Publications.
  12. Recombinant DNA by Watson, J.D.
  13. Essentials of Molecular Biology by Malacimski.
  14. Mobile DNA II by Nancy Craig, Martin Gellet Allan Lambowitz.

TH-XII ENVIRONMENTAL MICROBIAL TECHNOLGY

Unit – 1 Environment and Ecosystems
Definitions, biotic and abiotic environment. Environmental segments. Composition and structure of environment. Concept of biosphere, communities and ecosystems. Ecosystem characteristics, structure and function. Food chains, food webs and trophic structures. Ecological pyramids.

Unit – 2 Eutrophication
Water pollution and its control: Need for water management. Sources of water pollution. Measurement of water pollution, Eutrophication: Definition, causes of eutrophication, and microbial changes in eutrophic bodies of water induced by various inorganic pollutants. Effects of eutrophication on the quality of water environment, factors influencing eutrophication. Qualitative characteristics and properties of eutrophic lakes. Measurement of degree of eutrophication. Algae in eutrophication, algal blooms, their effects and toxicity, coloured waters, red tides, and cultural eutrophication. Physico-chemical and biological measures to control eutrophication.

Unit –3 Effluent treatment techniques
Microbiology of wastewater and solid waste treatment: – Waste-types-solid and liquid waste characterization, physical, chemical, biological, aerobic, anaerobic, primary, secondary and tertiary treatments.
Anaerobic processes: Anaerobic digestion, anaerobic filters, and upflow anaerobic sludge. Treatment schemes for effluents of dairy, distillery, tannery, sugar and antibiotic industries (Types, microbes used, types of Effluent Treatment Plants). Bioconversion of Solid Waste and utilization as fertilizer. Bioaccumulation of heavy metal ions from industrial effluents.

Unit – 4 Bioremediation of Xenobiotics
Microbiology of degradation of xenobiotics in the environment, ecological considerations, decay behaviour, biomagnification and degredative plasmids, hydrocarbons, substituted hydrocarbons, oil pollution, surfactants and pesticides. Genetically Modified Organisms released and its
environmental impact assessment and ethical issues.

Unit – 5 Global environmental problems
Ozone depletion, UV-B, green house effect and acid rain, their impact and biotechnological approaches for management. . Containment of acid mine drainage applying biomining [with reference to copper extraction from low grade ores].

References-

  1. Bioremediation by Baker K.H. And Herson D.S. 1994. MacGraw Hill Inc. N.Y.
  2. Waste Water Engineering – Treatment, Disposal and Re-use by Metcalf and Eddy, Inc., Tata MacGraw Hill, New Delhi.
  3. Pollution: Ecology and Biotreatment by Ec Eldowney, S. Hardman D.J. and Waite S.
  4. Longman Scientific Technical.
  5. Environmental Microbiology edited by Ralph Mitchell. A John Wiley and Sons. Inc.
  6. Waste Water Microbiology 2nd Edition by Bitton.
  7. Chemistry and Ecotoxicology of pollution. Edited by Des. W. Connell, G.J. Miller. Wiley Interscience Publications.
  8. Environmental Biotechnology. Edited by C. F. Forster and D.A., John Wase. Ellis Horwood Ltd. Publication.
  9. Advances in Waste Water Treatment Technologies. 1998. Volumes II and I by R. K. Trivedy. Global Science Publication.
  10. Biocatalysis and Biodegradation: Microbial transformation of organic compounds. 2000 by Lawrence P. Wacekett, C. Douglas Hershberger. ASM Publications.
  11. A Manual of Environmental Microbiology. 2nd Edition. 2001 by Christon J. Hurst (Chief Editor), ASM Publications.
  12. Biodegradation and Bioremediation, Academic Press, San Diego.
  13. Biotechnology in the sustainable environment, Plenum Press, N.Y.
  14. Basic Principles of Geomicrobiology by A. D. Agate, Pune.