• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of pnasPNASInfo for AuthorsSubscriptionsAboutThis Article
Proc Natl Acad Sci U S A. Jan 1, 1993; 90(1): 337–341.

Bovine pancreatic trypsin inhibitor-trypsin complex as a detection system for recombinant proteins.


Bovine pancreatic trypsin inhibitor (BPTI) binds to trypsin and anhydrotrypsin (an enzymatically inactive derivative of trypsin) with affinities of 6 x 10(-14) and 1.1 x 10(-13) M, respectively. We have taken advantage of the high affinity and specificity of this binding reaction to develop a protein tagging system in which biotinylated trypsin or biotinylated anhydrotrypsin is used as the reagent to detect recombinant fusion proteins into which BPTI has been inserted. Two proteins, opsin and growth hormone, were used as targets for insertional mutagenesis with BPTI. In each case, both domains of the fusion protein appear to be correctly folded. The fusion proteins can be specifically and efficiently detected by biotinylated trypsin or biotinylated anhydrotrypsin, as demonstrated by staining of transfected cells, protein blotting, affinity purification, and a mobility shift assay in SDS/polyacrylamide gels.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (1.6M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Kolodziej PA, Young RA. Epitope tagging and protein surveillance. Methods Enzymol. 1991;194:508–519. [PubMed]
  • Vincent JP, Lazdunski M. Trypsin-pancreatic trypsin inhibitor association. Dynamics of the interaction and role of disulfide bridges. Biochemistry. 1972 Aug 1;11(16):2967–2977. [PubMed]
  • Vincent JP, Peron-Renner M, Pudles J, Lazdunski M. The association of anhydrotrypsin with the pancreatic trypsin inhibitors. Biochemistry. 1974 Sep 24;13(20):4205–4211. [PubMed]
  • Levilliers N, Péron M, Arrio B, Pudles J. On the mechanism of action of proteolytic inhibitors. IV. Effect of 8 M urea on the stability of trypsin in trypsin-inhibitor complexes. Arch Biochem Biophys. 1970 Oct;140(2):474–483. [PubMed]
  • Huber R, Kukla D, Bode W, Schwager P, Bartels K, Deisenhofer J, Steigemann W. Structure of the complex formed by bovine trypsin and bovine pancreatic trypsin inhibitor. II. Crystallographic refinement at 1.9 A resolution. J Mol Biol. 1974 Oct 15;89(1):73–101. [PubMed]
  • Nathans J, Hogness DS. Isolation, sequence analysis, and intron-exon arrangement of the gene encoding bovine rhodopsin. Cell. 1983 Oct;34(3):807–814. [PubMed]
  • Leung DW, Spencer SA, Cachianes G, Hammonds RG, Collins C, Henzel WJ, Barnard R, Waters MJ, Wood WI. Growth hormone receptor and serum binding protein: purification, cloning and expression. Nature. 1987 Dec 10;330(6148):537–543. [PubMed]
  • DeNoto FM, Moore DD, Goodman HM. Human growth hormone DNA sequence and mRNA structure: possible alternative splicing. Nucleic Acids Res. 1981 Aug 11;9(15):3719–3730. [PMC free article] [PubMed]
  • Goldenberg DP. Kinetic analysis of the folding and unfolding of a mutant form of bovine pancreatic trypsin inhibitor lacking the cysteine-14 and -38 thiols. Biochemistry. 1988 Apr 5;27(7):2481–2489. [PubMed]
  • Ako H, Foster RJ, Ryan CA. The preparation of anhydro-trypsin and its reactivity with naturally occurring proteinase inhibitors. Biochem Biophys Res Commun. 1972 Jun 28;47(6):1402–1407. [PubMed]
  • Nathans J. Determinants of visual pigment absorbance: role of charged amino acids in the putative transmembrane segments. Biochemistry. 1990 Jan 30;29(4):937–942. [PubMed]
  • Hodges RS, Heaton RJ, Parker JM, Molday L, Molday RS. Antigen-antibody interaction. Synthetic peptides define linear antigenic determinants recognized by monoclonal antibodies directed to the cytoplasmic carboxyl terminus of rhodopsin. J Biol Chem. 1988 Aug 25;263(24):11768–11775. [PubMed]
  • de Vos AM, Ultsch M, Kossiakoff AA. Human growth hormone and extracellular domain of its receptor: crystal structure of the complex. Science. 1992 Jan 17;255(5042):306–312. [PubMed]
  • Cunningham BC, Wells JA. High-resolution epitope mapping of hGH-receptor interactions by alanine-scanning mutagenesis. Science. 1989 Jun 2;244(4908):1081–1085. [PubMed]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences


Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...


  • Compound
    PubChem Compound links
  • MedGen
    Related information in MedGen
  • PubMed
    PubMed citations for these articles
  • Substance
    PubChem Substance links