Recombinant Protein Techniques

  • Ian M. Rosenberg


The isolation and characterization of proteins parallels the major technical advances in protein chemistry. The first proteins that were characterized were those that could be isolated in large quantities. With the availability of more sensitive techniques, smaller and smaller quantities were required to produce detailed structural information. Newly evolving technologies, in conjunction with expression systems, have allowed the production of large amounts of any proteinacious factor. The isolation and structural characterization of proteins now depends only on the ingenuity of the investigator in devising an appropriate assay.


Fusion Protein Recombinant Protein Insect Cell Translation Product Wheat Germ Extract 
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  1. Andersons D, Engström Å, Josephson S, Hansson L, Steiner H (1991): Biologically active and amidated cecropin produced in a baculovirus expression system from a fusion construct containing the antibody-binding part of protein A. Biochem J 280: 219–224Google Scholar
  2. Andreason GL, Evans GA (1989): Optimization of electroporation for transfection of mammalian cell lines. Anal Biochem 180: 269–275CrossRefGoogle Scholar
  3. Ausubel FM, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, Struhl K (1993): Current Protocols in Molecular Biology. New York: John WileyGoogle Scholar
  4. Becker DM, Guarente L (1991): High efficiency transformation of yeast by e1ectroporation. Methods Enzymo1194:182–187Google Scholar
  5. Bennett MK, Garc’a-Arrarâs JE, Elferink LA, Peterson K, Fleming AM, Hazuka CD, Scheller RH (1993): The syntaxin family of vesicular transport receptors. Cell 74: 863–873CrossRefGoogle Scholar
  6. Brizzard BL, Chubet RG, Vizard DL (1994): Immunoaffinity purification of FLAG epitope-tagged bacterial alkaline phosphatase using a novel monoclonal antibody and peptide elution. Bio Techniques 16: 730–735Google Scholar
  7. Broker M (1994): Isolation of recombinant proteins from Saccharomyces cerevisiae by use of osmotically fragile mutant strains. Bio Techniques 16: 604–610Google Scholar
  8. Bush GL, Tassin A-M, Fridn H, Meyer DI (1991): Secretion in yeast. J Biol Chem 266: 13811–13814Google Scholar
  9. Dieckmann CL, Tzagoloff A (1985): Assembly of the mitochondria] membrane system. J Biol Chem 260: 1513–1520Google Scholar
  10. Fasman GD (1989): Prediction of Protein Structure and the Principles of Protein Conformation, Fasman GD, ed. New York: Plenum PressGoogle Scholar
  11. Feigner PL, Gadek TR, Holm M, Roman R, Chan HW, Wenz M, Northrop JP, Rinhold GM, Danielsen M (1987): Lipofection: A highly efficient, lipid mediated DNA transfection procedure. Proc Nail Acad Sci USA 84: 7413–7417CrossRefGoogle Scholar
  12. Field J, Nikawa J-I, Broek D, MacDonald B, Rodgers L, Wilson IA, Lerner RA, Wigler M (1988): Purification of a RAS-responsive adenylyl cyclase complex from Succharomyces cerevisiae by use of an epitope addition method. Mol Cell Biol 8: 2159 2165Google Scholar
  13. Frankel SR, Sohn R, Leinwand L (1991): The use of sarkosyl in generating soluble protein after bacterial expression. Proc Natl Acad Sci USA 88: 1192–1196CrossRefGoogle Scholar
  14. Gearing DP, Nicola NA, Metcalf D, Foote S, Willson TA, Gough NM, Williams RL (1989): Production of leukemia factor in E. coli by a novel procedure and its use in maintaining embryonic stem cells in culture. Bio/Technology 7: 1157–1161Google Scholar
  15. Gluzman Y (1981): SV-40 transformed simian cells support the replication of early 5V-40 mutants. Cell 23: 175–182CrossRefGoogle Scholar
  16. Green N, Alexander H, Olson A, Alexander S, Shinnick TM, Sutcliffe JG, Lerner RA (1982): Immunogenic structure of the influenza virus hemagglutinin. Cell 28: 477487Google Scholar
  17. Grieco F, Hay JM, Hull R (1992): An improved procedure for the purification of protein fused with glutathione S-transferase. Bio Techniques 13: 856–857Google Scholar
  18. Hochuli E, Bannwarth W, Döbeli H, Gentz R, Stüber D (1988): Genetic approach to facilitate purification of recombinant proteins with a novel chelate adsorbent. Bio/Technology 6: 1321–1325CrossRefGoogle Scholar
  19. Hodgson J (1993): Expression systems: a user’s guide. Bio/Technology 11:887–893 Holzman LB, Marks RM, Dixit VM (1990): A novel immediate-early response gene of endothelium is induced by cytokines and encodes a secreted protein. Mol Cell Biol 10: 5830–5838Google Scholar
  20. Hopp TP, Prickett KS, Price VL, Libby RT, March CJ, Cerretti DP, Urdal DL, Conlon PJ (1988): A short polypeptide marker sequence for recombinant protein identification and purification. Bio/Technology 6: 1204–1210CrossRefGoogle Scholar
  21. Ito H, Fukuda Y, Murata K, Kimura A (1983): Transformation of intact yeast cells treated with alkali cations. J Bacteriol 153: 163–168Google Scholar
  22. Kellermann OK, Ferenci T (1982): Maltose-binding protein from Escherichia coli. Methods Enzymol 90: 459–463Google Scholar
  23. Kim J-S, Raines RT (1993): Ribonuclease 5-peptide as a carrier in fusion proteins. Protein Science 2: 348–356CrossRefGoogle Scholar
  24. Kimmel AR, Berger SL, eds. (1987): Guide to Molecular Cloning. Methods in Enzymology Series, Vol 152. New York: Academic PressGoogle Scholar
  25. Klekamp MS, Weil PA (1982): Specific transcription of homologous class III genes in yeast Saccharomyces cerevisiae soluble cell-free extracts. J Biol Chem 257: 84328441Google Scholar
  26. Kunz D, Gerard NP, Gerard C (1992): The human leukocyte platelet-activating factor receptor. J Biol Chem 267: 9101–9106Google Scholar
  27. Lin K, Cheng S (1991): An efficient method to purify active eukaryotic proteins from the inclusion bodies in Escherichia coli. BioTechniques 11: 748–753Google Scholar
  28. Mak P, McDonnell DR Weigel NL, Schrader WT, O’Malley BW (1989): Expression of functional chicken oviduct progesterone receptors in yeast (Saccharomyces cerevisiae). J Biol Chem 264: 21613–21618Google Scholar
  29. Maniatis T, Fritsch EF, Sambrook J (1982): Molecular Cloning. Cold Spring Harbor, NY: Cold Spring Harbor LaboratoryGoogle Scholar
  30. Melton DA, Krieg PA, Rebagliati MR, Maniatis T, Zinn K, Green MR (1984): Efficient in vitro synthesis of biologically active RNA and RNA hybridization probes from plasmids containing a bacteriophage SPG promoter. Nucl Acids Res 12: 7057–7070CrossRefGoogle Scholar
  31. Morrow KJ (1995): Optimizing expression systems for the enhancement of protein purification. Gen Eng News May 1: 16–17Google Scholar
  32. Muzyczka N, ed. (1989): Eukaryotic viral expression vectors: Current Topics in Microbiology and Immunology. Berlin: Springer-VerlagGoogle Scholar
  33. Nagai K, Thogersen HC (1987): Synthesis and sequence-specific proteolysis of hybrid proteins produced in Escherichia coli. Methods Enzymol 153: 461–481CrossRefGoogle Scholar
  34. Nilsson B, Abrahmsén L (1990): Fusions to staphylococcal protein A. Methods Enzymol 185: 144–161CrossRefGoogle Scholar
  35. Nilsson B, Abrahmsén L, Uhlén M (1985): Immobilization and purification of enzymes with staphylococcal protein A gene fusion vectors. EMBO J 4: 1075–1080Google Scholar
  36. Nonet ML, Grundahl K, Meyer BJ, Rand JB (1993): Synaptic function is impared but not eliminated in C. elegans mutants lacking synaptotagmin. Cell 73: 1291–1305CrossRefGoogle Scholar
  37. Nuoffer C, Davidson HW, Matteson J, Meinkoth J, Balch WE (1994): A GDP-bound form of Rabi inhibits protein export from the endoplasmic reticulum and transport between golgi compartments. J Cell Bio 1125: 225–237CrossRefGoogle Scholar
  38. Nygren PA, Stähl S, Uhlén M (1994): Engineering proteins to facilitate bioprocessing. Trends Biotechnol 12: 338–349CrossRefGoogle Scholar
  39. Richards FM, Wyckoff HW (1971): Bovine pancreatic ribonuclease. In: The Enzymes, Vol 4, Boyer PD, ed. New York: Academic PressGoogle Scholar
  40. Rüther U, Müller-Hill B (1983): Easy identification of cDNA clones. EMBO J 2: 1791–1794Google Scholar
  41. Schmidt TG, Skerra A (1993): The random peptide library-assisted engineering of a C-terminal affinity peptide, useful for the detection and purification of a functional Ig Fv fragment. Protein Eng 6: 109–122CrossRefGoogle Scholar
  42. Smith DB, Johnson KS (1988): Single-step purification of polypeptides expressed in E. coli as fusions with glutathione S-transferase. Gene 67: 31–40Google Scholar
  43. Sorger PK, Pelham HRB (1987): Purification and characterization of a heat-shock element binding protein from yeast. EMBO J 6: 3035–3041Google Scholar
  44. Spindler KR, Rosser DSE, Berk AJ (1984): Analysis of Adenovirus transforming proteins from early regions lA and lB with antisera to inducible fusion antigens produced in Escherichia col. J Viro149: 132–141Google Scholar
  45. Su X, Prestwood AK, McGraw RA (1992): Production of recombinant porcine tumor necrosis factor alpha in a novel E. coli expression system. Bio Techniques 13: 756762Google Scholar
  46. Summers MD, Smith GE (1987): A Manual of Methods for Baculovirus Vectors and Insect Cell Culture Procedures. Texas Agricultural Experiment Station Bulletin No. 1555, College Station, TXGoogle Scholar
  47. Toneguzzo F, Keating A, Flynn S, McDonald K (1988): Electric field-mediated gene transfer: characterization of DNA transfer and patterns of integration in lymphoid cells. Nucleic Acids Res 16: 5515–5532CrossRefGoogle Scholar
  48. Wilson IA, Niman HL, Houghten RA, Cherenson AR, Connolly ML, Lerner RA (1984): The structure of an antigenic deteminant in a protein. Cell 37: 767–778CrossRefGoogle Scholar
  49. Wingfield PT, Mattaliano RJ, MacDonald HR, Craig S, Clove GM, Gronenborn AM, Schmiesner U (1987): Recombinant derived IL–la, replacing a labile asparagine with a serine. Protein Eng 1:413–27–36Google Scholar
  50. Wong TK, Neumann E (1982): Electric field mediated gene transfer. Biochem Biophys Res Commun 107: 584–587CrossRefGoogle Scholar

General References

  1. Goeddel DV, ed. (1990): Gene Expression Technology. Methods in Enzymology Series, Vol 185. New York: Academic PressGoogle Scholar
  2. Grossman L, Wu R, eds. (1987): Recombinant DNA, Parts D, E, F. Methods in Enzymology Series, Vols 153, 154, 155. New York: Academic PressGoogle Scholar
  3. Seetharam R, Sharma SK, eds. (1991): Purification and Analysis of Recombinant Proteins. New York: Marcel Dekker, Inc.Google Scholar
  4. Wu R, ed. (1992–1993): Recombinant DNA, Parts G, H, I. Methods in Enzymology Series, Vols 216, 217, 218. New York: Academic PressGoogle Scholar

Copyright information

© Springer Science+Business Media New York 1996

Authors and Affiliations

  • Ian M. Rosenberg
    • 1
  1. 1.Massachusetts General HospitalBostonUSA

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