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. 1975 Apr;147(1):139–144. doi: 10.1042/bj1470139

Isolation and characterization of a variant of ovomucoid.

A Waheed, A Salhuddin
PMCID: PMC1165383  PMID: 1171685

Abstract

A simple procedure, which can be used on a preparative scale, for the isolation and purification of a major variant of ovomucoid from egg white is described. Ovomucoid was precipitated by salt, and further fractionated by chromatography on sulphoethyl-Sephadex. It showed size homogeneity as revealed by gel chromatography and sodium dodecyl sulphate-polyacrylamide-gel electrophoresis where the mobility was consistent with a molecular weight of 28 300+/-2300. The inhibitor showed full antiryptic but no antichymotryptic activity. The u.v.-absorption and fluorescence characteristics indicated the absence of tryptophan. Polyacrylamide-gel electrophoresis in the presence of 9M-urea demonstrated absence of charge heterogeneity. The intrinsic viscosity of ovomucoid was 5.36ml/g which yielded an equivalent hydrodynamic radius (2.9nm), axial ratio (6.0) and frictional ratio (1.31) of the molecule. The Stokes radius (3.5nm), diffusion coefficient (7.8 times 10(-7 cm2/s) and frictional ratio (1.35) were calculated from gel-filtration data. These results suggest that ovomucoid exists in non-globular conformation under native conditions and that the deviation from the behaviour of a typical globular protein seems to be due both to asymmetry and hydration.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Ackers G. K. Analytical gel chromatography of proteins. Adv Protein Chem. 1970;24:343–446. doi: 10.1016/s0065-3233(08)60245-4. [DOI] [PubMed] [Google Scholar]
  2. Ahmad F., Salahuddin A. Influence of temperature on the intrinsic viscosities of proteins in random coil conformation. Biochemistry. 1974 Jan 15;13(2):245–249. doi: 10.1021/bi00699a003. [DOI] [PubMed] [Google Scholar]
  3. Ansari A. A., Salahuddin A. Purification and some properties of rabbit anti-ovalbumin. Biochem J. 1973 Dec;135(4):705–711. doi: 10.1042/bj1350705. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Beeley J. G. The isolation of ovomucoid variants differing in carbohydrate composition. Biochem J. 1971 Jul;123(3):399–405. doi: 10.1042/bj1230399. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. DAVIS B. J. DISC ELECTROPHORESIS. II. METHOD AND APPLICATION TO HUMAN SERUM PROTEINS. Ann N Y Acad Sci. 1964 Dec 28;121:404–427. doi: 10.1111/j.1749-6632.1964.tb14213.x. [DOI] [PubMed] [Google Scholar]
  6. Davis J. G., Mapes C. J., Donovan J. W. Batch purification of ovomucoid and characterization of the purified product. Biochemistry. 1971 Jan 5;10(1):39–42. doi: 10.1021/bi00777a006. [DOI] [PubMed] [Google Scholar]
  7. Donovan J. W. A spectrophotometric and spectrofluorometric study of intramolecular interactions of phenolic groups in ovomucoid. Biochemistry. 1967 Dec;6(12):3918–3927. doi: 10.1021/bi00864a038. [DOI] [PubMed] [Google Scholar]
  8. Duncombe W. G. The colorimetric micro-determination of long-chain fatty acids. Biochem J. 1963 Jul;88(1):7–10. doi: 10.1042/bj0880007. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. EYLAR E. H., JEANLOZ R. W. Oligosaccharides from the alpha1 acid glycoprotein of human plasma. J Biol Chem. 1962 Mar;237:622–628. [PubMed] [Google Scholar]
  10. FEENEY R. E., STEVENS F. C., OSUGA D. T. The specificities of chicken ovomucoid and ovoinhibitor. J Biol Chem. 1963 Apr;238:1415–1418. [PubMed] [Google Scholar]
  11. FRANCOIS C., MARSHALL R. D., NEUBERGER A. Carbohydrates in protein. 4. The determination of mannose in hen's-egg albumin by radioisotope dilution. Biochem J. 1962 May;83:335–341. doi: 10.1042/bj0830335. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. FREDERICQ E., DEUTSCH H. F. Studies on ovomucoid. J Biol Chem. 1949 Dec;181(2):499–510. [PubMed] [Google Scholar]
  13. GREEN N. M. Competition among trypsin inhibitors. J Biol Chem. 1953 Dec;205(2):535–551. [PubMed] [Google Scholar]
  14. Herskovits T. T., Jaillet H. Structural stability and solvent denaturation of myoglobin. Science. 1969 Jan 17;163(3864):282–285. doi: 10.1126/science.163.3864.282. [DOI] [PubMed] [Google Scholar]
  15. Kay E., Strickland E. H., Billups C. Near ultraviolet circular dichroism and absorption spectra of chicken ovomucoid and acetylated derivatives at 297 and 77 degrees K. J Biol Chem. 1974 Feb 10;249(3):797–802. [PubMed] [Google Scholar]
  16. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  17. Melamed M. D. Electrophoretic properties of ovomucoid. Biochem J. 1967 Jun;103(3):805–810. doi: 10.1042/bj1030805. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Narita K. End group determination. Mol Biol Biochem Biophys. 1970;8:25–90. doi: 10.1007/978-3-662-12834-3_3. [DOI] [PubMed] [Google Scholar]
  19. Osuga D. T., Feeney R. E. Biochemistry of the egg-white proteins of the ratite group. Arch Biochem Biophys. 1968 Mar 20;124(1):560–574. doi: 10.1016/0003-9861(68)90367-6. [DOI] [PubMed] [Google Scholar]
  20. PENASSE L., JUTISZ M., FROMAGEOT C., FRAENKEL-CONRAT H. La détermination des groupes carboxyliques des protéines. II. Le groupe carboxylique terminal de l'ovomucoide. Biochim Biophys Acta. 1952 Nov;9(5):551–556. doi: 10.1016/0006-3002(52)90205-9. [DOI] [PubMed] [Google Scholar]
  21. Parish C. R., Marchalonis J. J. A simple and rapid acrylamide gel method for estimating the molecular weights of proteins and protein subunits. Anal Biochem. 1970 Apr;34(2):436–450. doi: 10.1016/0003-2697(70)90128-4. [DOI] [PubMed] [Google Scholar]
  22. Pflumm M. N., Wang J. L., Edelman G. M. Conformational changes in concanavalin A. J Biol Chem. 1971 Jul 10;246(13):4369–4370. [PubMed] [Google Scholar]
  23. REISSIG J. L., STORMINGER J. L., LELOIR L. F. A modified colorimetric method for the estimation of N-acetylamino sugars. J Biol Chem. 1955 Dec;217(2):959–966. [PubMed] [Google Scholar]
  24. RHODES M. B., AZARI P. R., FEENEY R. E. Analysis, fractionation, and purification of egg white proteins with cellulose-cation exchanger. J Biol Chem. 1958 Jan;230(1):399–408. [PubMed] [Google Scholar]
  25. SVENNERHOLM L. Quantitative estimation of sialic acids. II. A colorimetric resorcinol-hydrochloric acid method. Biochim Biophys Acta. 1957 Jun;24(3):604–611. doi: 10.1016/0006-3002(57)90254-8. [DOI] [PubMed] [Google Scholar]
  26. WEIL L., TIMASHEFF S. N. On the inhibition of chymotrypsin by ovomucoid. Arch Biochem Biophys. 1960 Mar;87:134–136. doi: 10.1016/0003-9861(60)90134-x. [DOI] [PubMed] [Google Scholar]
  27. Weber K., Osborn M. The reliability of molecular weight determinations by dodecyl sulfate-polyacrylamide gel electrophoresis. J Biol Chem. 1969 Aug 25;244(16):4406–4412. [PubMed] [Google Scholar]

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