(Hyper)thermophilic Enzymes: Production and Purification

P. Falcicchio, M. Levisson, S.W.M. Kengen, S. Koutsopoulos

Research output: Chapter in Book/Report/Conference proceedingChapterAcademicpeer-review

4 Citations (Scopus)


The discovery of thermophilic and hyperthermophilic microorganisms, thriving at environmental temperatures near or above 100 °C, has revolutionized our ideas about the upper temperature limit at which life can exist. The characterization of (hyper)thermostable proteins has broadened our understanding and presented new opportunities for solving one of the most challenging problems in biophysics: how is structural stability and biological function maintained at high temperatures where “normal” proteins undergo dramatic structural changes? In our laboratory we have purified and studied many thermostable and hyperthermostable proteins in an attempt to determine the molecular basis of heat stability. Here, we present methods to express such proteins and enzymes in E. coli and provide a general protocol for overproduction and purification. The ability to produce enzymes that retain their stability and activity at elevated temperatures creates exciting opportunities for a wide range of biocatalytic applications
Original languageEnglish
Title of host publicationProtein Downstream Processing
EditorsN.E. Labrou
Place of PublicationTotowa
PublisherHumana Press
ISBN (Electronic)9781627039772
ISBN (Print)9781627039765
Publication statusPublished - 2014

Publication series

NameMethods in molecular biology
PublisherSpringer Verlag
ISSN (Print)1064-3745


  • Biocatalysis
  • Heterologous production
  • His-tag
  • Immobilized metal affinity chromatography (IMAC)
  • Protein purification
  • Size exclusion chromatography (SEC)
  • Thermal stability
  • Thermozymes


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