1887

Abstract

The gene encoding a pectate lyase from the strain sp. BP-23 was cloned and expressed in . The nucleotide sequence of a 1214 bp DNA fragment containing gene was determined, revealing an ORF of 666 nucleotides that encoded a protein of 23233 Da. The deduced amino acid sequence of the encoded enzyme showed homology to pectate lyases A, B, C and D from , Pel-3 and PelB from and PelI from . Homology was also found to the protein deduced from the gene, the function of which is unknown. The heterologous expressed enzyme depolymerized polygalacturonate and pectins of methyl esterification degree from 22 to 89%, and exhibited similar activity on polygalacturonate and on 89% esterified citrus pectin. Optimum temperature and pH for enzymic activity were 50 °C and pH 10, respectively. Ca was required for activity on pectic substrates, while the enzyme was strongly inhibited by Ba.

Keyword(s): lyase , pectate and pectin
Loading

Article metrics loading...

/content/journal/micro/10.1099/00221287-146-1-89
2000-01-01
2021-10-27
Loading full text...

Full text loading...

/deliver/fulltext/micro/146/1/1460089a.html?itemId=/content/journal/micro/10.1099/00221287-146-1-89&mimeType=html&fmt=ahah

References

  1. Alkorta I., Garbisu C., Llama M. J., Serra J. L. 1998; Industrial applications of pectic enzymes: a review. Process Biochem 33:21–28 [CrossRef]
    [Google Scholar]
  2. Altschul S. F., Madden T. L., Schaffer A. A., Zhang J., Zhang Z., Miller W., Lipman D. J. 1997; Gapped blast and psi-blast: a new generation of protein database search programs. Nucleic Acids Res 25:3389–3402 [CrossRef]
    [Google Scholar]
  3. Barras F., van Gijsegem F., Chatterjee A. K. 1994; Extracellular enzymes and pathogenesis of soft-rot Erwinia. Annu Rev Phytopathol 32:201–234 [CrossRef]
    [Google Scholar]
  4. Blanco A., Pastor F. I. J. 1993; Characterization of cellulase-free xylanases from the newly isolated Bacillus sp. strain BP-23. Can J Microbiol 39:1162–1166 [CrossRef]
    [Google Scholar]
  5. Blanco A., Vidal T., Colom J. F., Pastor F. I. J. 1995; Purification and properties of xylanase A from alkali-tolerant Bacillus sp. strain BP-23. Appl Environ Microbiol 61:4468–4470
    [Google Scholar]
  6. Blanco A., Dı́az P., Martı́nez J., Vidal T., Torres A. L., Pastor F. I. J. 1998; Cloning of a new endoglucanase gene from Bacillus sp. BP-23 and characterisation of the enzyme. Performance in paper manufacture from cereal straw. Appl Microbiol Biotechnol 50:48–54 [CrossRef]
    [Google Scholar]
  7. Blanco A., Dı́az P., Zueco J., Parascandola P., Pastor F. I. J. 1999; A multidomain xylanase from a Bacillus sp. with a region homologous to thermostabilizing domains of thermophilic enzymes. Microbiology 145:2163–2170 [CrossRef]
    [Google Scholar]
  8. Brühlmann F., Keen N. T. 1997; Cloning, sequence and expression of the pel gene from an Amycolata sp. Gene 202:45–51 [CrossRef]
    [Google Scholar]
  9. Burns J. K. 1991; The polygalacturonases and lyases. In The Chemistry and Technology of Pectin pp. 165–188Edited by Walter R. H. San Diego: Academic Press;
    [Google Scholar]
  10. Charkowski A. O., Alfano J. R., Preston G., Yuan J., He S. Y., Collmer A. 1998; The Pseudomonas syringae pv. tomato HrpW protein has domains similar to harpins and pectate lyases and can elicit the plant hypersensitive response and bind to pectate. J Bacteriol 180:5211–5217
    [Google Scholar]
  11. Collmer A., Ried J. L., Mount M. S. 1988; Assay methods for pectic enzymes. Methods Enzymol 161:329–335
    [Google Scholar]
  12. Dubnau D., Davidoff-Abelson R. 1971; Fate of transforming DNA following uptake by competent Bacillus subtilis. I. Formation and properties of the donor–recipient complex. J Mol Biol 56:209–221 [CrossRef]
    [Google Scholar]
  13. Godessart N., Muñoa F. J., Regue M., Juárez A. 1988; Chromosomal mutations that increase the production of a plasmid-encoded haemolysin in Escherichia coli. J Gen Microbiol 134:2779–2787
    [Google Scholar]
  14. González-Candelas L., Kolattukudy P. E. 1992; Isolation and analysis of a novel inducible pectate lyase gene from the phytopathogenic fungus Fusarium solani f. sp. pisi (Nectria haematococca, mating population VI). J Bacteriol 174:6343–6349
    [Google Scholar]
  15. Guo W., González-Candelas L., Kolattukudy P. E. 1995a; Cloning of a new pectate lyase gene pelC from Fusarium solani f. sp. pisi (Nectria haematococca, mating type VI) and characterization of the gene product expressed in Pichia pastoris. Arch Biochem Biophys 323:352–360 [CrossRef]
    [Google Scholar]
  16. Guo W., González-Candelas L., Kolattukudy P. E. 1995b; Cloning of a novel constitutively expressed pectate lyase gene pelB from Fusarium solani f. sp. pisi (Nectria haematococca, mating type VI) and characterization of the gene product expressed in Pichia pastoris. J Bacteriol 177:7070–7077
    [Google Scholar]
  17. Guo W., González-Candelas L., Kolattukudy P. E. 1996; Identification of a novel pelD gene expressed uniquely in planta by Fusarium solani f. sp. pisi (Nectria haematococca, mating type VI) and characterization of its protein product as an endo-pectate lyase. Arch Biochem Biophys 332:305–312 [CrossRef]
    [Google Scholar]
  18. Hanahan D. 1983; Studies on transformation of Escherichia coli with plasmids. J Mol Biol 166:557–580 [CrossRef]
    [Google Scholar]
  19. von Heijne G. 1986; A new method for predicting signal sequence cleavage sites. Nucleic Acids Res 14:4683–4690 [CrossRef]
    [Google Scholar]
  20. Heikinheimo R., Flego D., Pirhonen M., Karlsson M. B., Eriksson A., Mäe A., Köiv V., Palva E. T. 1995; Characterization of a novel pectate lyase from Erwinia carotovora subsp. carotovora. Mol Plant–Microbe Interact 8:207–217 [CrossRef]
    [Google Scholar]
  21. Henriksson G., Akin D. E., Slomczynski D., Eriksson K.-E. L. 1999; Production of highly efficient enzymes for flax retting by Rhizomucor pusillus. J Biotechnol 68:115–123 [CrossRef]
    [Google Scholar]
  22. Henrissat B., Heffron S. E., Yoder M. D., Lietzke S. E., Jurnak F. 1995; Functional implications of structure-based sequence alignment of proteins in the extracellular pectate lyase superfamily. Plant Physiol 107:963–976 [CrossRef]
    [Google Scholar]
  23. Hinton J. C. D., Sidebotham J. M., Gill D. R., Salmond G. P. C. 1989; Extracellular and periplasmic isoenzymes of pectate lyase from Erwinia carotovora subspecies carotovora belong to different gene families. Mol Microbiol 3:1785–1795 [CrossRef]
    [Google Scholar]
  24. Hugouvieux-Cotte-Pattat N., Condemine G., Nasser W., Reverchon S. 1996; Regulation of pectinolysis in Erwinia chrysanthemi. Annu Rev Microbiol 50:213–257 [CrossRef]
    [Google Scholar]
  25. Keen N. T., Dahlbeck D., Staskawicz B., Belser W. 1984; Molecular cloning of pectate lyase genes from Erwinia chrysanthemi and their expression in Escherichia coli. J Bacteriol 159:825–831
    [Google Scholar]
  26. Kim J. F., Beer S. V. 1998; HrpW of Erwinia amylovora, a new harpin that contains a domain homologous to pectate lyases of a distinct class. J Bacteriol 180:5203–5210
    [Google Scholar]
  27. Kobayashi T., Koike K., Yoshimatsu T., Higaki N., Suzumatsu A., Ozawa T., Hatada Y., Ito S. 1999; Purification and properties of a low-molecular-weight, high-alkaline pectate lyase from an alkaliphilic strain of Bacillus. Biosci Biotechnol Biochem 63:65–72 [CrossRef]
    [Google Scholar]
  28. Kozianowski G., Canganella F., Rainey F. A., Hippe H., Antranikian G. 1997; Purification and characterization of thermostable pectate-lyases from a newly isolated thermophilic bacterium, Thermoanaerobacter italicus sp. nov. Extremophiles 1:171–182 [CrossRef]
    [Google Scholar]
  29. Kunst F., Ogasawara N., Moszer I.148 other authors 1997; The complete genome sequence of the gram-positive bacterium. Bacillus subtilis. Nature 390:249–256 [CrossRef]
    [Google Scholar]
  30. Laemmli U. K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685 [CrossRef]
    [Google Scholar]
  31. Liu Y., Chatterjee A., Chatterjee A. K. 1994; Nucleotide sequence and expression of a novel pectate lyase gene (pel-3) and a closely linked endopolygalacturonase gene (peh-1) of Erwinia carotovora subsp. carotovora 71. Appl Environ Microbiol 60:2545–2552
    [Google Scholar]
  32. Nagarajan V. 1993; Protein secretion. In Bacillus subtilis and Other Gram-positive Bacteria: Biochemistry, Physiology, and Molecular Genetics pp. 713–726Edited by Sonenshein A. L., Hoch J. A., Losick R. Washington, DC: American Society for Microbiology;
    [Google Scholar]
  33. Nasser W., Chalet F., Robert-Baudouy J. 1990; Purification and characterization of extracellular pectate lyase from Bacillus subtilis. Biochimie 72:689–695 [CrossRef]
    [Google Scholar]
  34. Nasser W., Awadé A. C., Reverchon S., Robert-Baudouy J. 1993; Pectate lyase from Bacillus subtilis: molecular characterization of the gene, and properties of the cloned enzyme. FEBS Lett 335:319–326 [CrossRef]
    [Google Scholar]
  35. Sakamoto T., Hours R. A., Sakai T. 1994; Purification, characterization, and production ot two pectic transeliminases with protopectinase activity from Bacillus subtilis. Biosci Biotechnol Biochem 58:353–358 [CrossRef]
    [Google Scholar]
  36. Sakamoto T., Kawasaki H., Sakai T. 1996; Molecular cloning and nucleotide sequence of the gene encoding phosphate-inducible pectin lyase of Bacillus subtilis. FEBS Lett 398:269–273 [CrossRef]
    [Google Scholar]
  37. Sambrook J., Fritsch E. F., Maniatis T. 1989 Molecular Cloning: a Laboratory Manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  38. Shevchik V. E., Robert-Baudouy J., Hugouvieux-Cotte-Pattat N. 1997; Pectate lyase PelI of Erwinia chrysanthemi 3937 belongs to a new family. J Bacteriol 179:7321–7330
    [Google Scholar]
  39. Shevchik V. E., Boccara M., Vedel R., Hugouvieux-Cotte-Pattat N. 1998; Processing of the pectate lyase PelI by extracellular proteases of Erwinia chrysanthemi 3937. Mol Microbiol 29:1459–1469 [CrossRef]
    [Google Scholar]
  40. Tardy F., Nasser W., Robert-Baudouy J., Hugouvieux-Cotte-Pattat N. 1997; Comparative analysis of the five major Erwinia chrysanthemi pectate lyases: enzyme characteristics and potential inhibitors. J Bacteriol 179:2503–2511
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/00221287-146-1-89
Loading
/content/journal/micro/10.1099/00221287-146-1-89
Loading

Data & Media loading...

Most cited this month Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error