PMID: 9183009May 1, 1997Paper

Molecular cloning and nucleotide sequence of an endo-1,5-alpha-L-arabinase gene from Bacillus subtilis

European Journal of Biochemistry
T SakamotoT Sakai

Abstract

The nucleotide sequence of the gene encoding an endo-1,5-alpha-L-arabinase (protopectinase C) of Bacillus subtilis was determined by sequencing fragments amplified by the cassette-ligation-mediated PCR (CLM-PCR). The gene covering the start and stop codon was amplified by PCR with two specific primers, which were designed from the sequence data determined by CLM-PCR. An approximately 1.5-kb amplification product was cloned into the vector pUC119, forming a plasmid termed pPPC. An ORF that encodes the arabinase composed of 324 amino acids including a 33-amino-acid signal peptide was assigned. Comparison of the deduced amino acid sequence of the enzyme with that of an Aspergillus niger endoarabinase showed 37% identity in a 207-amino-acid overlap. The optimal nucleotide sequence for catabolite repression of B. subtilis was found upstream of the structural gene. In a culture of Escherichia coli DH5alpha cells harboring pPPC, no arabinase activity was detected, either intracellularly or extracellularly, suggesting that the B. subtilis promotor is not functional in this transformant. In B. subtilis IFO 3134 strain, production of protopectinase C was repressed by readily metabolizable carbohydrates. In contrast, productivity (total e...Continue Reading

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Citations

Dec 25, 2010·Applied Microbiology and Biotechnology·Tatsuji SakamotoNaoya Kasai
Dec 4, 2003·Biochimica Et Biophysica Acta·T SakamotoH Kawasaki
Aug 26, 2000·Biochemical Engineering Journal·T MatsumotoH Fukuda
Feb 20, 2004·Journal of Bacteriology·Maria Paiva RaposoIsabel de Sá-Nogueira
Oct 6, 2000·Annual Review of Microbiology·J Stülke, W Hillen
Nov 24, 2004·FEMS Microbiology Letters·Teresa Fontes Leal, Isabel de Sá-Nogueira
May 20, 2000·Biotechnology & Genetic Engineering Reviews·R A PradeA J Mort
Jan 15, 2015·Journal of Agricultural and Food Chemistry·Zhou ChenZhengqiang Jiang

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