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Degradation of rice bran hemicellulose by Paenibacillus sp. strain HC1: gene cloning, characterization and function of β-D-glucosidase as an enzyme involved in degradation

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Abstract

A bacterium (strain HC1) capable of assimilating rice bran hemicellulose was isolated from a soil and identified as belonging to the genus Paenibacillus through taxonomical and 16S rDNA sequence analysis. Strain HC1 cells grown on rice bran hemicellulose as a sole carbon source inducibly produced extracellular xylanase and intracellular glycosidases such as β-d-glucosidase and β-d-arabinosidase. One of them, β-d-glucosidase was further analyzed. A genomic DNA library of the bacterium was constructed in Escherichia coli and gene coding for β-d-glucosidase was cloned by screening for β-d-glucoside-degrading phenotype in E. coli cells. Nucleotide sequence determination indicated that the gene for the enzyme contained an open reading frame consisting of 1,347 bp coding for a polypeptide with a molecular mass of 51.4 kDa. The polypeptide exhibits significant homology with other bacterial β-d-glucosidases and belongs to glycoside hydrolase family 1. β-d-Glucosidase purified from E. coli cells was a monomeric enzyme with a molecular mass of 50 kDa most active at around pH 7.0 and 37°C. Strain HC1 glycosidases responsible for degradation of rice bran hemicellulose are expected to be useful for structurally determining and molecularly modifying rice bran hemicellulose and its derivatives.

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Abbreviations

RB:

Rice bran

TLC:

Thin-layer chromatography

X-Glc:

5-Bromo-4-chrolo-3-indoryl-β-d-glucopyranoside

p-NP:

p-nitrophenyl

HPLC:

High-performance liquid chromatography

KPB:

Potassium phosphate buffer

SDS-PAGE:

Sodium dodecyl sulfate-polyacrylamide gel electrophoresis

aa:

Amino acids

acc no:

Accession number

bglA :

Strain HC1 β-d-glucosidase gene

BglA:

Strain HC1 β-d-glucosidase

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Acknowledgements

This work was supported in part by Grants-in-Aid from the Ministry of Education, Culture, Sports, Science and Technology of Japan to K.M. and W.H. and by the Program of Basic Research Activities for Innovative Biosciences (PROBRAIN) of Japan to K.M. A part of this work was also supported by the Iijima foundation of Japan.

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Authors and Affiliations

  1. Laboratory of Basic and Applied Molecular Biotechnology, Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Uji, Kyoto, 611-0011, Japan

    Karen Mine Harada, Wataru Hashimoto & Kousaku Murata

  2. Chukyo Community College, Mizunami, Gifu, 509-6101, Japan

    Keiko Tanaka & Yasuki Fukuda

Authors
  1. Karen Mine Harada
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  2. Keiko Tanaka
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  3. Yasuki Fukuda
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  4. Wataru Hashimoto
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  5. Kousaku Murata
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Correspondence to Kousaku Murata.

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Harada, K.M., Tanaka, K., Fukuda, Y. et al. Degradation of rice bran hemicellulose by Paenibacillus sp. strain HC1: gene cloning, characterization and function of β-D-glucosidase as an enzyme involved in degradation. Arch Microbiol 184, 215–224 (2005). https://doi.org/10.1007/s00203-005-0038-8

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  • DOI: https://doi.org/10.1007/s00203-005-0038-8

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