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Cloning and sequencing of cel5Z gene from Erwinia chrysanthemi PY35.
Mol Cells. 2000 Jun 30; 10(3):269-74.MC

Abstract

The phytopathogenic bacterium Erwinia chrysanthemi (Ech) secretes multiple isozymes of plant cell wall disrupting enzymes such as pectate lyase and endoglucanases. We cloned genomic DNA from Ech PY35 digested with Sau3AI and ligated into pBluescript II SK+. One of the E. coli XL1-blue clones had the ability to hydrolyze carboxymethyl cellulose and polygalacturonic acid. By subsequent subcloning from this 2.9 kb fragment, we obtained a 2.0 kb (pPY401), designated cel5Z, which had the activity of hydrolyzation of carboxymethyl cellulose. The cel5Z gene had an open reading frame (ORF) of 1,281 bp starting with an ATG start codon and followed by a TAA stop codon, encoding 426 amino acids with a signal peptide of 41 amino acids. Since the deduced amino acid sequence of this protein was very similar to that of CelE of Pseudomonas fluorescens, and had the conserved region, VIYEIYNEPL, it belonged to the glycoside hydrolase family 5 of EC 3.2.1.4. The molecular mass of Cel5Z protein from E. coli XL1-blue, as analyzed by CMC-SDS-PAGE, appeared to be 42 kDa. The optimum pH was 6, and the optimum temperature was about 40 degrees C for its enzymatic activity.

Authors+Show Affiliations

Department of Agricultural Chemistry, Gyeongsang National University, Chinju, Korea.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

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

Language

eng

PubMed ID

10901164

Citation

Park, S R., et al. "Cloning and Sequencing of cel5Z Gene From Erwinia Chrysanthemi PY35." Molecules and Cells, vol. 10, no. 3, 2000, pp. 269-74.
Park SR, Kim MK, Kim JO, et al. Cloning and sequencing of cel5Z gene from Erwinia chrysanthemi PY35. Mol Cells. 2000;10(3):269-74.
Park, S. R., Kim, M. K., Kim, J. O., Cho, S. J., Cho, Y. U., & Yun, H. D. (2000). Cloning and sequencing of cel5Z gene from Erwinia chrysanthemi PY35. Molecules and Cells, 10(3), 269-74.
Park SR, et al. Cloning and Sequencing of cel5Z Gene From Erwinia Chrysanthemi PY35. Mol Cells. 2000 Jun 30;10(3):269-74. PubMed PMID: 10901164.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Cloning and sequencing of cel5Z gene from Erwinia chrysanthemi PY35. AU - Park,S R, AU - Kim,M K, AU - Kim,J O, AU - Cho,S J, AU - Cho,Y U, AU - Yun,H D, PY - 2000/7/20/pubmed PY - 2001/2/28/medline PY - 2000/7/20/entrez SP - 269 EP - 74 JF - Molecules and cells JO - Mol Cells VL - 10 IS - 3 N2 - The phytopathogenic bacterium Erwinia chrysanthemi (Ech) secretes multiple isozymes of plant cell wall disrupting enzymes such as pectate lyase and endoglucanases. We cloned genomic DNA from Ech PY35 digested with Sau3AI and ligated into pBluescript II SK+. One of the E. coli XL1-blue clones had the ability to hydrolyze carboxymethyl cellulose and polygalacturonic acid. By subsequent subcloning from this 2.9 kb fragment, we obtained a 2.0 kb (pPY401), designated cel5Z, which had the activity of hydrolyzation of carboxymethyl cellulose. The cel5Z gene had an open reading frame (ORF) of 1,281 bp starting with an ATG start codon and followed by a TAA stop codon, encoding 426 amino acids with a signal peptide of 41 amino acids. Since the deduced amino acid sequence of this protein was very similar to that of CelE of Pseudomonas fluorescens, and had the conserved region, VIYEIYNEPL, it belonged to the glycoside hydrolase family 5 of EC 3.2.1.4. The molecular mass of Cel5Z protein from E. coli XL1-blue, as analyzed by CMC-SDS-PAGE, appeared to be 42 kDa. The optimum pH was 6, and the optimum temperature was about 40 degrees C for its enzymatic activity. SN - 1016-8478 UR - https://www.unboundmedicine.com/medline/citation/10901164/Cloning_and_sequencing_of_cel5Z_gene_from_Erwinia_chrysanthemi_PY35_ L2 - http://www.molcells.org/journal/view.html?year=2000&volume=10&number=3&spage=269 DB - PRIME DP - Unbound Medicine ER -