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. 1990 Dec;56(12):3735–3740. doi: 10.1128/aem.56.12.3735-3740.1990

Production, Purification, and Properties of Extracellular Carboxyl Esterases from Bacillus subtilis NRRL 365

K Meghji 1, O P Ward 1,*, A Araujo 1,†
PMCID: PMC185060  PMID: 16348375

Abstract

Bacillus subtilis NRRL 365 produced high extracellular carboxyl esterase activity in submerged culture media containing wheat bran, corn steep liquor, and salts. Supplementation of this medium with glucose reduced esterase activity to 37% of that in the unsupplemented control. Esterase activity was purified by ammonium sulfate fractionation, DEAE-Sephadex A-50 ion-exchange chromatography with sodium chloride gradient elution, and preparative polyacrylamide gel electrophoresis. The resultant purified components, esterases I and II, manifested single bands following silver staining of polyacrylamide gel electrophoresis gels and had final specific activities of 80 and 520 U/mg, respectively. Molecular weights for components I and II were 36,000 and 105,000 to 110,000, respectively. Esterases I and II both had a pH optimum of 8.0, with relative activities of 10 and 85%, respectively, at pH 9.0. Kms with p-nitrophenylacetate were 0.91 mM for esterase I and 0.67 mM for esterase II. In general, patterns of enzyme inhibition were similar for both components. Differences were observed in the relative activities of esterases I and II towards p-nitrophenyl esters of acetate, propionate, and butyrate; Activity ratios for components I and II were 100:94:48 and 100:36:23, respectively. The purified components did not hydrolyze long-chain triglycerides and did not manifest proteolytic activity.

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Selected References

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

  1. 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]
  2. Higerd T. B. Isolation of acetyl esterase mutants of Bacillus subtilis 168. J Bacteriol. 1977 Feb;129(2):973–977. doi: 10.1128/jb.129.2.973-977.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Higerd T. B., Spizizen J. Isolation of two acetyl esterases from extracts of Bacillus subtilis. J Bacteriol. 1973 Jun;114(3):1184–1192. doi: 10.1128/jb.114.3.1184-1192.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Kennedy M. B., Lennarz W. J. Characterization of the extracellular lipase of Bacillus subtilis and its relationship to a membrane-bound lipase found in a mutant strain. J Biol Chem. 1979 Feb 25;254(4):1080–1089. [PubMed] [Google Scholar]
  5. 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]
  6. Matsunaga A., Koyama N., Noso Y. Purification and properties of esterase from Bacillus stearothermophilus. Arch Biochem Biophys. 1974 Feb;160(2):504–513. doi: 10.1016/0003-9861(74)90427-5. [DOI] [PubMed] [Google Scholar]
  7. Merril C. R., Goldman D., Sedman S. A., Ebert M. H. Ultrasensitive stain for proteins in polyacrylamide gels shows regional variation in cerebrospinal fluid proteins. Science. 1981 Mar 27;211(4489):1437–1438. doi: 10.1126/science.6162199. [DOI] [PubMed] [Google Scholar]
  8. Nakagawa A., Tsujita T., Okuda H. Purification and some properties of intracellular esterase from Pseudomonas fluorescens. J Biochem. 1984 Apr;95(4):1047–1054. doi: 10.1093/oxfordjournals.jbchem.a134692. [DOI] [PubMed] [Google Scholar]
  9. ORNSTEIN L. DISC ELECTROPHORESIS. I. BACKGROUND AND THEORY. Ann N Y Acad Sci. 1964 Dec 28;121:321–349. doi: 10.1111/j.1749-6632.1964.tb14207.x. [DOI] [PubMed] [Google Scholar]
  10. Parkkinen E. Multiple forms of carboxylesterases in Baker's yeast. Cell Mol Biol Incl Cyto Enzymol. 1980;26(2):147–154. [PubMed] [Google Scholar]
  11. Riefler J. F., 3rd, Higerd T. B. Characterization of intracellular esterase A from Bacillus subtilis. Biochim Biophys Acta. 1976 Mar 11;429(1):191–197. doi: 10.1016/0005-2744(76)90041-3. [DOI] [PubMed] [Google Scholar]
  12. Shum A. C., Markovetz A. J. Purification and properties of undecyl acetate esterase from Pseudomonas cepacia grown on 2-tridecanone. J Bacteriol. 1974 Jun;118(3):880–889. doi: 10.1128/jb.118.3.880-889.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Shum A. C., Markovetz A. J. Specificity and induction of undecyl acetate esterase from Pseudomonas cepacia grown on 2-tridecanone. J Bacteriol. 1974 Jun;118(3):890–897. doi: 10.1128/jb.118.3.890-897.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Sierra G. Dissociatiion of esterase from proteinase activity of Bacillus subtilis spores. Can J Microbiol. 1967 Jun;13(6):673–678. doi: 10.1139/m67-089. [DOI] [PubMed] [Google Scholar]
  15. Zaks A., Klibanov A. M. Enzymatic catalysis in organic media at 100 degrees C. Science. 1984 Jun 15;224(4654):1249–1251. doi: 10.1126/science.6729453. [DOI] [PubMed] [Google Scholar]

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