Skip to main content
NCBI home page
Biochemical Journal logoLink to Biochemical Journal
. 1997 Jun 1;324(Pt 2):403–411. doi: 10.1042/bj3240403

Two genes encoding an endoglucanase and a cellulose-binding protein are clustered and co-regulated by a TTA codon in Streptomyces halstedii JM8.

A L Garda 1, J M Fernández-Abalos 1, P Sánchez 1, A Ruiz-Arribas 1, R I Santamaría 1
PMCID: PMC1218445  PMID: 9182697

Abstract

Streptomyces halstedii JM8 Cel2 is an endoglucanase of 28 kDa that is first produced as a protein of 42 kDa (p42) and is later processed at its C-terminus. Cel2 displays optimal activity towards CM-cellulose at pH6 and 50 degrees C and shows no activity against crystalline cellulose or xylan. The N-terminus of p42 shares similarity with cellulases included in family 12 of the beta-glycanases and the C-terminus shares similarity with bacterial cellulose-binding domains included in family II. This latter domain enables the precursor to bind so tightly to Avicel that it can only be eluted by boiling in 10% (w/v) SDS. Another open reading frame (ORF) situated 216 bp downstream from the p42 ORF encodes a protein of 40 kDa (p40) that does not have any clear hydrolytic activity against cellulosic or xylanosic compounds, but shows high affinity for Avicel (crystalline cellulose). The p40 protein is processed in old cultures to give a protein of 35 kDa that does not bind to Avicel. Translation of both ORFs is impaired in Streptomyces coelicolor bldA mutants, suggesting that a TTA codon situated at the fourth position of the first ORF is responsible for this regulation. S1 nuclease protection experiments demonstrate that both ORFs are co-transcribed.

Full Text

The Full Text of this article is available as a PDF (839.4 KB).

Selected References

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

  1. BERNFELD P. Enzymes of starch degradation and synthesis. Adv Enzymol Relat Subj Biochem. 1951;12:379–428. doi: 10.1002/9780470122570.ch7. [DOI] [PubMed] [Google Scholar]
  2. Beall J. A., Mitchell G. F. Identification of a particular antigen from a parasite cDNA library using antibodies affinity purified from selected portions of Western blots. J Immunol Methods. 1986 Feb 12;86(2):217–223. doi: 10.1016/0022-1759(86)90456-4. [DOI] [PubMed] [Google Scholar]
  3. Chater K. F., Hopwood D. A., Kieser T., Thompson C. J. Gene cloning in Streptomyces. Curr Top Microbiol Immunol. 1982;96:69–95. doi: 10.1007/978-3-642-68315-2_5. [DOI] [PubMed] [Google Scholar]
  4. Din N., Forsythe I. J., Burtnick L. D., Gilkes N. R., Miller R. C., Jr, Warren R. A., Kilburn D. G. The cellulose-binding domain of endoglucanase A (CenA) from Cellulomonas fimi: evidence for the involvement of tryptophan residues in binding. Mol Microbiol. 1994 Feb;11(4):747–755. doi: 10.1111/j.1365-2958.1994.tb00352.x. [DOI] [PubMed] [Google Scholar]
  5. Favaloro J., Treisman R., Kamen R. Transcription maps of polyoma virus-specific RNA: analysis by two-dimensional nuclease S1 gel mapping. Methods Enzymol. 1980;65(1):718–749. doi: 10.1016/s0076-6879(80)65070-8. [DOI] [PubMed] [Google Scholar]
  6. Fernández-Abalos J. M., Sánchez P., Coll P. M., Villanueva J. R., Pérez P., Santamaría R. I. Cloning and nucleotide sequence of celA1, and endo-beta-1,4-glucanase-encoding gene from Streptomyces halstedii JM8. J Bacteriol. 1992 Oct;174(20):6368–6376. doi: 10.1128/jb.174.20.6368-6376.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Fernández-Moreno M. A., Caballero J. L., Hopwood D. A., Malpartida F. The act cluster contains regulatory and antibiotic export genes, direct targets for translational control by the bldA tRNA gene of Streptomyces. Cell. 1991 Aug 23;66(4):769–780. doi: 10.1016/0092-8674(91)90120-n. [DOI] [PubMed] [Google Scholar]
  8. Garda-Salas A. L., Santamaria R. I., Marcos M. J., Zhadan G. G., Villar E., Shnyrov V. L. Differential scanning calorimetry of the irreversible thermal denaturation of cellulase from Streptomyces halstedii JM8. Biochem Mol Biol Int. 1996 Feb;38(1):161–170. [PubMed] [Google Scholar]
  9. Gramajo H. C., Takano E., Bibb M. J. Stationary-phase production of the antibiotic actinorhodin in Streptomyces coelicolor A3(2) is transcriptionally regulated. Mol Microbiol. 1993 Mar;7(6):837–845. doi: 10.1111/j.1365-2958.1993.tb01174.x. [DOI] [PubMed] [Google Scholar]
  10. Hamamoto T., Foong F., Shoseyov O., Doi R. H. Analysis of functional domains of endoglucanases from Clostridium cellulovorans by gene cloning, nucleotide sequencing and chimeric protein construction. Mol Gen Genet. 1992 Feb;231(3):472–479. doi: 10.1007/BF00292718. [DOI] [PubMed] [Google Scholar]
  11. Hansen C. K., Diderichsen B., Jørgensen P. L. celA from Bacillus lautus PL236 encodes a novel cellulose-binding endo-beta-1,4-glucanase. J Bacteriol. 1992 Jun;174(11):3522–3531. doi: 10.1128/jb.174.11.3522-3531.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Henikoff S. Unidirectional digestion with exonuclease III creates targeted breakpoints for DNA sequencing. Gene. 1984 Jun;28(3):351–359. doi: 10.1016/0378-1119(84)90153-7. [DOI] [PubMed] [Google Scholar]
  13. Henrissat B. A classification of glycosyl hydrolases based on amino acid sequence similarities. Biochem J. 1991 Dec 1;280(Pt 2):309–316. doi: 10.1042/bj2800309. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Henrissat B., Bairoch A. New families in the classification of glycosyl hydrolases based on amino acid sequence similarities. Biochem J. 1993 Aug 1;293(Pt 3):781–788. doi: 10.1042/bj2930781. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Higgins D. G., Sharp P. M. CLUSTAL: a package for performing multiple sequence alignment on a microcomputer. Gene. 1988 Dec 15;73(1):237–244. doi: 10.1016/0378-1119(88)90330-7. [DOI] [PubMed] [Google Scholar]
  16. Igo M. M., Losick R. Regulation of a promoter that is utilized by minor forms of RNA polymerase holoenzyme in Bacillus subtilis. J Mol Biol. 1986 Oct 20;191(4):615–624. doi: 10.1016/0022-2836(86)90449-3. [DOI] [PubMed] [Google Scholar]
  17. Ishaque M., Kluepfel D. Cellulase complex of a mesophilic Streptomyces strain. Can J Microbiol. 1980 Feb;26(2):183–189. doi: 10.1139/m80-028. [DOI] [PubMed] [Google Scholar]
  18. Jung E. D., Lao G., Irwin D., Barr B. K., Benjamin A., Wilson D. B. DNA sequences and expression in Streptomyces lividans of an exoglucanase gene and an endoglucanase gene from Thermomonospora fusca. Appl Environ Microbiol. 1993 Sep;59(9):3032–3043. doi: 10.1128/aem.59.9.3032-3043.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  20. Lao G., Ghangas G. S., Jung E. D., Wilson D. B. DNA sequences of three beta-1,4-endoglucanase genes from Thermomonospora fusca. J Bacteriol. 1991 Jun;173(11):3397–3407. doi: 10.1128/jb.173.11.3397-3407.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Leskiw B. K., Lawlor E. J., Fernandez-Abalos J. M., Chater K. F. TTA codons in some genes prevent their expression in a class of developmental, antibiotic-negative, Streptomyces mutants. Proc Natl Acad Sci U S A. 1991 Mar 15;88(6):2461–2465. doi: 10.1073/pnas.88.6.2461. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Leskiw B. K., Mah R., Lawlor E. J., Chater K. F. Accumulation of bldA-specified tRNA is temporally regulated in Streptomyces coelicolor A3(2). J Bacteriol. 1993 Apr;175(7):1995–2005. doi: 10.1128/jb.175.7.1995-2005.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Marck C. 'DNA Strider': a 'C' program for the fast analysis of DNA and protein sequences on the Apple Macintosh family of computers. Nucleic Acids Res. 1988 Mar 11;16(5):1829–1836. doi: 10.1093/nar/16.5.1829. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Millward-Sadler S. J., Poole D. M., Henrissat B., Hazlewood G. P., Clarke J. H., Gilbert H. J. Evidence for a general role for high-affinity non-catalytic cellulose binding domains in microbial plant cell wall hydrolases. Mol Microbiol. 1994 Jan;11(2):375–382. doi: 10.1111/j.1365-2958.1994.tb00317.x. [DOI] [PubMed] [Google Scholar]
  25. Morrissey J. H. Silver stain for proteins in polyacrylamide gels: a modified procedure with enhanced uniform sensitivity. Anal Biochem. 1981 Nov 1;117(2):307–310. doi: 10.1016/0003-2697(81)90783-1. [DOI] [PubMed] [Google Scholar]
  26. Murray M. G. Use of sodium trichloroacetate and mung bean nuclease to increase sensitivity and precision during transcript mapping. Anal Biochem. 1986 Oct;158(1):165–170. doi: 10.1016/0003-2697(86)90605-6. [DOI] [PubMed] [Google Scholar]
  27. O'Neill G., Goh S. H., Warren R. A., Kilburn D. G., Miller R. C., Jr Structure of the gene encoding the exoglucanase of Cellulomonas fimi. Gene. 1986;44(2-3):325–330. doi: 10.1016/0378-1119(86)90197-6. [DOI] [PubMed] [Google Scholar]
  28. Ooi T., Shinmyo A., Okada H., Hara S., Ikenaka T., Murao S., Arai M. Cloning and sequence analysis of a cDNA for cellulase (FI-CMCase) from Aspergillus aculeatus. Curr Genet. 1990 Oct;18(3):217–222. doi: 10.1007/BF00318384. [DOI] [PubMed] [Google Scholar]
  29. Pearson W. R., Lipman D. J. Improved tools for biological sequence comparison. Proc Natl Acad Sci U S A. 1988 Apr;85(8):2444–2448. doi: 10.1073/pnas.85.8.2444. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Perito B., Hanhart E., Irdani T., Iqbal M., McCarthy A. J., Mastromei G. Characterization and sequence analysis of a Streptomyces rochei A2 endoglucanase-encoding gene. Gene. 1994 Oct 11;148(1):119–124. doi: 10.1016/0378-1119(94)90244-5. [DOI] [PubMed] [Google Scholar]
  31. Piret J. M., Chater K. F. Phage-mediated cloning of bldA, a region involved in Streptomyces coelicolor morphological development, and its analysis by genetic complementation. J Bacteriol. 1985 Sep;163(3):965–972. doi: 10.1128/jb.163.3.965-972.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Poole D. M., Hazlewood G. P., Huskisson N. S., Virden R., Gilbert H. J. The role of conserved tryptophan residues in the interaction of a bacterial cellulose binding domain with its ligand. FEMS Microbiol Lett. 1993 Jan 1;106(1):77–83. doi: 10.1111/j.1574-6968.1993.tb05938.x. [DOI] [PubMed] [Google Scholar]
  33. Ruiz-Arribas A., Fernández-Abalos J. M., Sánchez P., Garda A. L., Santamariá R. I. Overproduction, purification, and biochemical characterization of a xylanase (Xys1) from Streptomyces halstedii JM8. Appl Environ Microbiol. 1995 Jun;61(6):2414–2419. doi: 10.1128/aem.61.6.2414-2419.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Saarilahti H. T., Henrissat B., Palva E. T. CelS: a novel endoglucanase identified from Erwinia carotovora subsp. carotovora. Gene. 1990 May 31;90(1):9–14. doi: 10.1016/0378-1119(90)90433-r. [DOI] [PubMed] [Google Scholar]
  35. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Schlochtermeier A., Walter S., Schröder J., Moorman M., Schrempf H. The gene encoding the cellulase (Avicelase) Cel1 from Streptomyces reticuli and analysis of protein domains. Mol Microbiol. 1992 Dec;6(23):3611–3621. doi: 10.1111/j.1365-2958.1992.tb01797.x. [DOI] [PubMed] [Google Scholar]
  37. Teather R. M., Wood P. J. Use of Congo red-polysaccharide interactions in enumeration and characterization of cellulolytic bacteria from the bovine rumen. Appl Environ Microbiol. 1982 Apr;43(4):777–780. doi: 10.1128/aem.43.4.777-780.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Théberge M., Lacaze P., Shareck F., Morosoli R., Kluepfel D. Purification and characterization of an endoglucanase from Streptomyces lividans 66 and DNA sequence of the gene. Appl Environ Microbiol. 1992 Mar;58(3):815–820. doi: 10.1128/aem.58.3.815-820.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Vieira J., Messing J. Production of single-stranded plasmid DNA. Methods Enzymol. 1987;153:3–11. doi: 10.1016/0076-6879(87)53044-0. [DOI] [PubMed] [Google Scholar]
  40. Wittmann S., Shareck F., Kluepfel D., Morosoli R. Purification and characterization of the CelB endoglucanase from Streptomyces lividans 66 and DNA sequence of the encoding gene. Appl Environ Microbiol. 1994 May;60(5):1701–1703. doi: 10.1128/aem.60.5.1701-1703.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Zhang S., Lao G., Wilson D. B. Characterization of a Thermomonospora fusca exocellulase. Biochemistry. 1995 Mar 14;34(10):3386–3395. doi: 10.1021/bi00010a030. [DOI] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES