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Journal of Bacteriology, November 2006, p. 7592-7599, Vol. 188, No. 21
0021-9193/06/$08.00+0     doi:10.1128/JB.00729-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Identification and Characterization of the Bacillus thuringiensis phaZ Gene, Encoding New Intracellular Poly-3-Hydroxybutyrate Depolymerase

Chi-Ling Tseng, Hui-Ju Chen, and Gwo-Chyuan Shaw^*

Institute of Biochemistry and Molecular Biology, School of Life Science, National Yang-Ming University, Taipei, Taiwan, Republic of China

Received 21 May 2006/ Accepted 16 August 2006

A gene that codes for a novel intracellular poly-3-hydroxybutyrate (PHB) depolymerase has now been identified in the genome of Bacillus thuringiensis subsp. israelensis ATCC 35646. This gene, previously annotated as a hypothetical 3-oxoadipate enol-lactonase (PcaD) gene and now designated phaZ, encodes a protein that shows no significant similarity with any known PHB depolymerase. Purified His-tagged PhaZ could efficiently degrade trypsin-activated native PHB granules as well as artificial amorphous PHB granules and release 3-hydroxybutyrate monomer as a hydrolytic product, but it could not hydrolyze denatured semicrystalline PHB. In contrast, purified His-tagged PcaD of Pseudomonas putida was unable to degrade trypsin-activated native PHB granules and artificial amorphous PHB granules. The B. thuringiensis PhaZ was inactive against p-nitrophenylpalmitate, tributyrin, and triolein. Sonication supernatants of the wild-type B. thuringiensis cells exhibited a PHB-hydrolyzing activity in vitro, whereas those prepared from a phaZ mutant lost this activity. The phaZ mutant showed a higher PHB content than the wild type at late stationary phase of growth in a nutrient-rich medium, indicating that this PhaZ can function as a PHB depolymerase in vivo. PhaZ contains a lipase box-like sequence (G-W-S₁₀₂-M-G) but lacks a signal peptide. A purified His-tagged S102A variant had lost the PHB-hydrolyzing activity. Taken together, these results indicate that B. thuringiensis harbors a new type of intracellular PHB depolymerase.

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* Corresponding author. Mailing address: Institute of Biochemistry and Molecular Biology, School of Life Science, National Yang-Ming University, Taipei 112, Taiwan. Phone: 886-2-2826-7127. Fax: 886-2-2826-4843. E-mail: gcshaw{at}ym.edu.tw.

Published ahead of print on 25 August 2006.

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Journal of Bacteriology, November 2006, p. 7592-7599, Vol. 188, No. 21
0021-9193/06/$08.00+0     doi:10.1128/JB.00729-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

This article has been cited by other articles:

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