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Molecular cloning and characterization of a cellulase gene from a symbiotic protist of the lower termite, Coptotermes formosanus.
Gene. 2005 Apr 11; 349:67-75.GENE

Abstract

The endo-beta-1,4-glucanase gene was cloned from a cDNA library constructed from the mixed population of symbiotic protists in the hindgut of the lower termite, Coptotermes formosanus, using the lambda ZAP II vector. The recombinant phage library was screened for cellulolytic activity by the Congo red staining procedure. The nucleotide sequence comprised 941 nucleotides including a polyA tail sequence and showed high sequence similarity with endoglucanase genes belonging to glycosyl hydrolase family 5. Determination of the 5' end of the cellulase gene using the 5'RACE method showed that the full-length cDNA comprised a 921-bp ORF, encoding a putative 33,620 Da protein. The organismal source of this cellulase gene was identified using PCR with gene-specific primers and whole-cell in situ hybridization as the smallest symbiotic hypermastigote protist, Spirotrichonympha leidyi. The optimal pH and temperature of the cellulase heterologously expressed in Escherichia coli were 5.8-6.0 and 70 degrees C, respectively. The Km and Vmax values on carboxymethyl cellulose (CMC) substrate were 1.90 mg/ml and 148.2 units/mg protein, respectively.

Authors+Show Affiliations

JST Bio-Recycle Project/Environmental Molecular Biology Laboratory, RIKEN, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan. i-tetsu@riken.jpNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Comparative Study
Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

15777663

Citation

Inoue, Tetsushi, et al. "Molecular Cloning and Characterization of a Cellulase Gene From a Symbiotic Protist of the Lower Termite, Coptotermes Formosanus." Gene, vol. 349, 2005, pp. 67-75.
Inoue T, Moriya S, Ohkuma M, et al. Molecular cloning and characterization of a cellulase gene from a symbiotic protist of the lower termite, Coptotermes formosanus. Gene. 2005;349:67-75.
Inoue, T., Moriya, S., Ohkuma, M., & Kudo, T. (2005). Molecular cloning and characterization of a cellulase gene from a symbiotic protist of the lower termite, Coptotermes formosanus. Gene, 349, 67-75.
Inoue T, et al. Molecular Cloning and Characterization of a Cellulase Gene From a Symbiotic Protist of the Lower Termite, Coptotermes Formosanus. Gene. 2005 Apr 11;349:67-75. PubMed PMID: 15777663.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Molecular cloning and characterization of a cellulase gene from a symbiotic protist of the lower termite, Coptotermes formosanus. AU - Inoue,Tetsushi, AU - Moriya,Shigeharu, AU - Ohkuma,Moriya, AU - Kudo,Toshiaki, PY - 2004/09/02/received PY - 2004/11/15/revised PY - 2004/11/26/accepted PY - 2005/3/22/pubmed PY - 2005/5/28/medline PY - 2005/3/22/entrez SP - 67 EP - 75 JF - Gene JO - Gene VL - 349 N2 - The endo-beta-1,4-glucanase gene was cloned from a cDNA library constructed from the mixed population of symbiotic protists in the hindgut of the lower termite, Coptotermes formosanus, using the lambda ZAP II vector. The recombinant phage library was screened for cellulolytic activity by the Congo red staining procedure. The nucleotide sequence comprised 941 nucleotides including a polyA tail sequence and showed high sequence similarity with endoglucanase genes belonging to glycosyl hydrolase family 5. Determination of the 5' end of the cellulase gene using the 5'RACE method showed that the full-length cDNA comprised a 921-bp ORF, encoding a putative 33,620 Da protein. The organismal source of this cellulase gene was identified using PCR with gene-specific primers and whole-cell in situ hybridization as the smallest symbiotic hypermastigote protist, Spirotrichonympha leidyi. The optimal pH and temperature of the cellulase heterologously expressed in Escherichia coli were 5.8-6.0 and 70 degrees C, respectively. The Km and Vmax values on carboxymethyl cellulose (CMC) substrate were 1.90 mg/ml and 148.2 units/mg protein, respectively. SN - 0378-1119 UR - https://www.unboundmedicine.com/medline/citation/15777663/Molecular_cloning_and_characterization_of_a_cellulase_gene_from_a_symbiotic_protist_of_the_lower_termite_Coptotermes_formosanus_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0378-1119(04)00724-3 DB - PRIME DP - Unbound Medicine ER -