[2] Preparation of myoglobins

https://doi.org/10.1016/0076-6879(81)76112-3Get rights and content

Publisher Summary

This chapter presents procedures for the isolation of intracellular oxygen-binding proteins of tissues, called “tissue hemoglobins” in the widest sense. All of these, except Ascaris and yeast hemoglobin, are monomers or dimers having a minimum molecular weight of 18,000 with similar optical spectra and chemical reactivity. Strictly, only muscle hemoglobin should be called “myoglobin”; by extension the term is often applied to other tissue hemoglobins as well. Ferric myoglobin may be purified by chromatography on carboxymethyl (CM) cellulose, usually at slightly acid pH or on diethylaminoethyl (DEAE) cellulose. The choice of preparative procedure depends on the use to which the purified myoglobin will be put. Both DEAE and CM ion-exchange columns yield myoglobin that is pure in the sense of being free from contaminating polypeptide chains. Better resolution of forms of myoglobin differing only in charge is achieved on CM-cellulose. Such columns, however, are usually operated at acid pH, and it is a matter of experience that oxymyoglobin exposed to mildly acidic conditions becomes ferric and, in the process, undergoes some minor but apparently irreversible change. The chapter also explains the isolation and purification of vertebrate myoglobins.

References (80)

  • M.Z. Atassi

    Biochim. Biophys. Acta

    (1970)
  • A. Akeson et al.

    Arch. Biochem. Biophys.

    (1960)
  • K.D. Hardman et al.

    J. Biol. Chem.

    (1966)
  • K.D. Hapner et al.

    J. Biol. Chem.

    (1968)
  • W.D. Brown

    J. Biol. Chem.

    (1961)
  • I. Yamazaki et al.

    J. Biol. Chem.

    (1964)
  • T.E. Hugli et al.

    J. Biol. Chem.

    (1970)
  • T. Gotoh et al.

    J. Chromatogr.

    (1971)
  • A.B. Edmundson et al.

    J. Mol. Biol.

    (1962)
  • T. Takano

    J. Mol. Biol.

    (1977)
  • T. Takano

    J. Mol. Biol.

    (1977)
  • H. Scouloudi et al.

    J. Mol. Biol.

    (1978)
  • A. Rossi Fanelli et al.

    Arch. Biochem. Biophys.

    (1955)
  • E.E. Lattman et al.

    J. Mol. Biol.

    (1971)
  • G.J. Fosmire et al.

    Comp. Biochem. Physiol.

    (1976)
  • R.H. Kretsinger

    J. Mol. Biol.

    (1968)
  • S. Schuder et al.

    Anal. Biochem.

    (1979)
  • D.J. Goss et al.

    J. Biochem. Biophys. Methods

    (1980)
  • A. Hayashi et al.

    Biochim. Biophys. Acta

    (1973)
  • W. Steigemann et al.

    J. Mol. Biol.

    (1979)
  • E. Weber et al.

    J. Mol. Biol.

    (1978)
  • T. Imamura et al.

    J. Biol. Chem.

    (1972)
  • S.N. Vinogradov et al.

    J. Biol. Chem.

    (1970)
  • B. Seamonds et al.

    J. Biol. Chem.

    (1971)
  • B. Seamonds et al.

    J. Biol. Chem.

    (1976)
  • E. Padlan et al.

    J. Biol. Chem.

    (1974)
  • M. Brunori et al.

    J. Mol. Biol.

    (1968)
  • M. Brunori et al.

    J. Mol. Biol.

    (1972)
  • G. Rotilio et al.

    Biochim. Biophys. Acta

    (1971)
  • B.A. Wittenberg et al.

    Arch. Biochem. Biophys.

    (1965)
  • T.L. Blundell et al.

    J. Mol. Biol.

    (1975)
  • C.P. Mangum
  • K.R.H. Read
  • A.S. Bonner et al.

    FEBS Lett.

    (1977)
  • T. Okazaki et al.

    Biochim. Biophys. Acta

    (1967)
  • J.B. Wittenberg et al.

    J. Biol. Chem.

    (1974)
  • M.H. Smith et al.

    Arch. Biochem. Biophys.

    (1962)
  • E. Steers et al.

    Comp. Biochem. Physiol.

    (1979)
  • M. J. Dilworth, this series, Vol. 69...
  • H. Theorell

    Biochem. Z.

    (1932)
  • Cited by (74)

    • X-ray fluorescence holography of biological metal sites: Application to myoglobin

      2022, Biochemical and Biophysical Research Communications
    • Functional inhibition of redox regulated heme proteins: A novel mechanism towards oxidative stress induced by homocysteine

      2021, Redox Biology
      Citation Excerpt :

      Preparation of protein stock solutions and determination of concentrations- Protein solutions were dialyzed extensively against 0.1 M KCl solution at 4 °C and filtered using Millipore syringe filter (0.22 μm). Concentrations of protein solutions were determined using ϵ, molar extinction coefficient values of 39000 M−1 cm−1 at 280 nm for Lyz [20], 9800 M−1cm−1 at 277.5 nm for RNase-A [21], 1,05,000 M−1cm−1 at 405 nm for HRP [22], 1,09,000 M−1cm−1 at 409 nm for Cyt c [23], 1,79,000 M−1cm−1 at 405 nm for Hb [24] and 1,16,000 at 408 nm for Myo [25]. All solutions for optical measurements were prepared in degassed 0.05 M potassium phosphate buffer (pH 7.4) containing 0.1 M KCl.

    • Long-term incubation of myoglobin with glyoxal induces amyloid like aggregation of the heme protein: Implications of advanced glycation end products in protein conformational disorders

      2021, Journal of Molecular Liquids
      Citation Excerpt :

      All other reagents were AR grade and purchased locally. Purchased Mb which is mostly in met, deoxy form, was dissolved in 50 mM potassium phosphate buffer (PB), pH 6.0, and concentration determined from its Soret absorbance using ε408 nm = 116 mM−1 cm−1 (monomer basis) [49,50,52–56]. Both Mb and glyoxal solutions were sterile-filtered and sterile conditions were maintained during incubation of the samples.

    • Methylglyoxal-induced modification causes aggregation of myoglobin

      2016, Spectrochimica Acta - Part A: Molecular and Biomolecular Spectroscopy
      Citation Excerpt :

      All other reagents were AR grade and purchased locally. Mb was dissolved in 50 mM potassium phosphate buffer (PB), pH 6.0, and concentration was determined considering ε408 nm = 116 mMˉ1 cmˉ1 [37]. The stock concentration of MG in water was estimated using ε284 nm = 12.3 Mˉ1 cmˉ1 [38].

    View all citing articles on Scopus
    View full text