Working with Enzymes

David Skrincosky1, Cindy Santangelo1

1 Worthington Biochemical, Lakewood, New Jersey
Publication Name:  Current Protocols Essential Laboratory Techniques
Unit Number:  Unit 10.1
DOI:  10.1002/9780470089941.et1001s00
Online Posting Date:  October, 2008
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Abstract

This unit briefly describes the history, theory, and biochemical principles of enzymes. The enzyme classification system, which is based upon the reactions enzymes catalyze, is explained. Several factors affecting enzyme activity including concentration, temperature, pH, and substrate choices are reviewed. While it is difficult to make generalizations since each enzyme is unique, we have compiled useful tips on enzyme storage and commonly encountered forms. This unit also applies the basic general principles of enzymes to a specific class of enzymes, called restriction endonucleases, in order to demonstrate how to correctly set up an enzymatic reaction.

Keywords: enzyme; units; classification; kinetics; inhibition; specificity; restriction endonucleases

     
 
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Table of Contents

  • Introduction
  • Overview of Enzymes
  • Handling Enzymes in the Laboratory
  • Example of Setting Up an Enzymatic Reaction: Restriction Enzymes
  • Literature Cited
  • Figures
  • Tables
     
 
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Materials

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Figures

Videos

Literature Cited

   Bloch, K.D. and Grossmann, B. 1995. Digestion of DNA with restriction endonucleases. Curr. Protoc. Mol. Biol. 31:3.1.1‐3.1.21.
   Burrell, M.M. (ed.) 1993. Enzymes of Molecular Biology. In Methods in Molecular Biology, Vol. 16. Humana Press, Totowa, N.J.
   Harlow, E. and Lane, D. 1998. Antibodies: A Laboratory Manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.
   Holum, J. 1968. Elements of General and Biological Chemistry, 2nd ed. pp. 377. John Wiley & Sons, New York.
   Mishra, N.C. 2002. Nucleases: Molecular Biology and Applications. John Wiley & Sons, Hoboken, NJ.
   Sambrook, J.and Russell, D. 2001. Molecular Cloning A Laboratory Manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.
   Stern, K. 1936. On the mechanism of enzyme action. J. Biol. Chem. 114:473‐494.
   Worthington, V. (ed.) 1993. Worthington Enzyme Manual. Worthington Biochemical, Lakewood, N.J.
Key References
   Barrett, A.J., Rawlings, N.D., and Woessner, J.F. 1998. Handbook of Proteolytic Enzymes. Academic Press, London.
  Detailed information on proteases.
   Burrell, M.M. (ed.) 1993. See above.
  Provides a more extensive review of restriction enzymes than necessary for this chapter.
   Coligan, J.E., Dunn, B.M., Speicher, D.W., and Wingfield, P.T. (eds.) 2003. Short Protocols in Protein Science. John Wiley & Sons, Hoboken, N.J.
  Includes protocols for common enzyme techniques.
   Copeland, R.A. 2000. Enzymes: A Practical Introduction to Structure Mechanism, and Data Analysis. Wiley‐VCH, New York.
  A comprehensive review of enzymology.
   Mishra, N.C. 2000. See above.
  Provides extensive reviews of restriction enzymes.
   Whitford, D. 2005. Proteins Structure and Function. John Wiley & Sons, London.
  An excellent reference for protein basics.
  Methods In Enzymology series. Academic Press.
  Highly respected series in life sciences publishing topics of current interest.
Internet Resources
  http://www.expasy.ch/enzyme
  Web site containing information relative to nomenclature of enzymes and links to individual enzymes. A description of the Web site can be found in Bairoch, A. 2000. The ENZYME database in 2000. Nucleic Acids Res 28:304‐305.
  http://www.chem.qmul.ac.uk/iubmb/enzyme
  Web site for enzyme nomenclature and links to details about specific enzymes. Contains information published in the book Enzyme Nomenclature (Academic Press), as well as in the European Journal of Biochemistry between 1994 and 1999 (details on the Web site).
  http://www.worthington‐biochem.com
  Web site containing characteristics and references for numerous enzymes.
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