Identification of an alkaline-tolerant cyclodextrin-metabolizing bacterium and characterization of its cyclodextrinase gene.
Bacillus circulans A11, an alkaline-tolerant cyclodextrin-metabolizing bacterium isolated from South-East Asian soil, was reidentified as Paenibacillus sp. A11 based on 16S rRNA gene sequence comparison, G + C content and cellular fatty acid composition. Levels of similarity of the 16S rRNA gene between strain A11 and the Paenibacillus species were 90-99%, while similarity with Bacillus circulans was only 86%. The major cellular fatty acid was anteiso-C15:0 which accounted for 59.3% of the total cellular fatty acids and the G+C content was 50.3 mol%. The CDase gene coding for this enzyme was cloned into E. coli. The open reading frame of the CDase gene was 1,959 bp encoding a CDase of 653 amino acid residues. At maximum growth, the specific activity of the recombinant CDase from E. coli was higher than that of Paenibacillus sp. A11. By SDS-PAGE, the translation product of the recombinant gene showed the same mobility as the purified CDase from the original strain. CDase from both Paenibacillus sp. A11 and E. coli produced glucose and maltose as dominant end-products of beta-CD hydrolysis. The ratio of maltose to glucose was 1:2.
Kaulpiboon
Jarunee
J
Biological Science Ph.D Program, Faculty of Science, Chulalongkorn University, Bangkok 10330, Thailand.
Rimphanitchayakit
Vichien
V
Pongsawasdi
Piamsook
P
D000595:Amino Acid Sequence; D001210:Asia, Southeastern; D001407:Bacillus / Q000145:classification* / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D001482:Base Composition; D001483:Base Sequence; D003001:Cloning, Molecular; D003505:Cyclodextrins / Q000378:metabolism*; D004269:DNA, Bacterial / Q000737:chemistry / Q000302:isolation & purification; D004275:DNA, Ribosomal / Q000737:chemistry / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005227:Fatty Acids / Q000032:analysis; D020459:Genes, rRNA; D005947:Glucose / Q000032:analysis; D006026:Glycoside Hydrolases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008320:Maltose / Q000032:analysis; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010802:Phylogeny; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017385:Sequence Homology; D012988:Soil Microbiology; D047392:beta-Cyclodextrins / Q000378:metabolism
D001426:Bacterial Proteins; D003505:Cyclodextrins; D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D005227:Fatty Acids; D012336:RNA, Ribosomal, 16S; D011994:Recombinant Proteins; D047392:beta-Cyclodextrins; D008320:Maltose; D006026:Glycoside Hydrolases; C048745:cyclomaltodextrinase; D005947:Glucose