Archives of Microbiology

, Volume 147, Issue 1, pp 64–67 | Cite as

Characterization of the amine oxidase involved in the growth of Trichosporon cutaneum X4 on ethylamine as source of carbon, nitrogen and energy

  • P. J. Large
  • L. A. Sherlock
Original Papers

Abstract

The amine oxidase from Trichosporon cutaneum X4 grown on ethylamine as carbon, nitrogen and energy source was purified to near homogeneity. The purified enzyme showed the highest resistance to heat of any amine oxidase hitherto characterized from a yeast (half-life at 62°C, 14 min). Measurement of kinetic parameters as a function of carbon chain length showed results typical of a benzylamine oxidase. Both non-denaturing- and sodium dodecyl sulphate-polyacrylamide gel electrophoresis showed multiple bands, and dimethyl suberimidate cross-linking studies revealed that the enzyme consisted of multimers of two polypeptide chains of Mr respectively 19,000 and 26,000. The smallest structure to show activity probably contained two of each kind of subunit.

Key words

Trichosporon cutaneum X4 Ethylamine Benzylamine oxidase Heat resistance Subunit composition Dimethyl suberimidate cross-linking 

Abbreviation

SDS

sodium dodecyl sulphate

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Andrews P (1970) Estimation of molecular size and molecular weights of biological compounds by gel filtration. Meth Biochem Anal 18:1–53Google Scholar
  2. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254Google Scholar
  3. Davies GE, Stark GR (1970) Use of dimethyl suberimidate, a cross-linking reagent, in studying the subunit structure of oligomeric proteins. Proc Natl Acad Sci USA 66:651–656Google Scholar
  4. Ferguson KA (1964) Starch-gel electrophoresis/application to the classification of pituitary proteins and polypeptides. Metabolism 13:985–1002Google Scholar
  5. Green J, Large PJ (1984) Regulation of the key enzymes of methylated amine metabolism in Candida boidinii. J Gen Microbiol 130:1947–1959Google Scholar
  6. Green J, Haywood GW, Large PJ (1983) Serological differences between the multiple amine oxidases of yeasts and comparison of the specificities of the purified enzymes from Candida utilis and Pichia pastoris. Biochem J 211:481–493Google Scholar
  7. Haywood GW, Large PJ (1981) Microbial oxidation of amines. Distribution, purification and properties of two primary amine oxidases from the yeast Candida boidinii grown on amines as sole nitrogen source. Biochem J 199:187–201Google Scholar
  8. Heath LA, Large PJ (1984) A single amine oxidase mediates utilization of primary aliphatic amines as nitrogen source by Kluyveromyces lactis. FEMS Microbiol Lett 22:15–19Google Scholar
  9. Hedrick JL, Smith AJ (1968) Size and charge isomer separation and estimation of molecular weights of proteins by disc gel electrophoresis. Arch Biochem Biophys 126:155–164Google Scholar
  10. Martin RG, Ames BN (1961) A method for determining the sedimentation behavior of enzymes: application to protein mixtures. J Biol Chem 236:1372–1379Google Scholar
  11. Middelhoven WJ, Hoogkamer-te Niet MC, Kreger-van Rij NJW (1984) Trichosporon adeninovorans sp. nov., a yeast species utilizing adenine, xanthine, uric acid, putrescine and primary n-alkylamines as sole source of carbon, nitrogen and energy. Antonie van Leeuwenhoek J Microbiol Serol 50:369–378Google Scholar
  12. Middelhoven WJ, de Kievit H, Biesbroek AL (1985) Yeast species utilizing uric acid, adenine, n-alkylamines and diamines as sole source of carbon and energy. Antonie van Leeuwenhoek J Microbiol Serol 51:289–301Google Scholar
  13. Sherlock LA, Large PJ (1985) Control of benzylamine oxidase activity in Kluyveromyces fragilis grown on primary amines as nitrogen source. FEMS Microbiol Lett 29:99–104Google Scholar
  14. Sherlock LA, Large PJ, Whitaker RG (1986) A new type of methylamine oxidase: the sole oxidase produced during growth of Sporobolomyces albo-rubescens on primary alkylamines. Yeast 2:87–92Google Scholar
  15. Van Dijken JP, Bos P (1981) Utilization of amines by yeasts. Arch Microbiol 128:320–324Google Scholar
  16. Veenhuis M, van der Klei IJ, Harder W (1986) Physiological role of microbodies in the yeast Trichosporon cutaneum during growth on ethylamine as the source of energy, carbon and nitrogen. Arch Microbiol 145:39–50Google Scholar
  17. Zwart KB, Harder W (1983) Regulation of the metabolism of some alkylated amines in the yeasts Candida utilis and Hansenula polymorpha. J Gen Microbiol 129:3157–3169Google Scholar

Copyright information

© Springer-Verlag 1987

Authors and Affiliations

  • P. J. Large
    • 1
  • L. A. Sherlock
    • 1
  1. 1.Department of BiochemistryUniversity of HullHullUK

Personalised recommendations