Identification of the 4-hydroxycinnamate decarboxylase (PAD) gene of Klebsiella oxytoca.
A 7.1-kbp DNA fragment isolated from a wild strain of Klebsiella oxytoca was sequenced, leading to the identification of 10 open-reading frames (ORFs), including a 504-bp Pad gene. The Pad gene of the Gram-negative bacterium was subsequently expressed in Escherichia coli as a chimeric Pad. The deduced amino acid (AA) sequence of the Pad gene from wild-type K. oxytoca showed approximately 50% homology to those of other bacterial PADs from Gram-positive bacilli plus a coccus. These data and a genomic library search of some gamma-proteobacteria, including E. coli and Vibrio sp., indicated that PAD of K. oxytoca is a member of the bacterial PAD family characteristic of Gram-negative bacteria. Using Pad-specific PCR primers designed from the Gram-negative bacterial Pad of K. oxytoca, Pad genes of two further strains of K. oxytoca, another wild isolate and JCM 1665 and two PAD-positive Enterobacter spp. were successfully amplified for specific Pad detection.
Uchiyama
Hirofumi
H
Division of Applied Bioscience, Research Faculty of Agriculture, Hokkaido University, Sapporo, Japan.
Hashidoko
Yasuyuki
Y
Kuriyama
Yuhki
Y
Tahara
Satoshi
S
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics; D002262:Carboxy-Lyases / Q000235:genetics*; D017124:Conserved Sequence; D017931:DNA Primers; D006090:Gram-Negative Bacteria / Q000201:enzymology / Q000235:genetics; D006094:Gram-Positive Bacteria / Q000201:enzymology / Q000235:genetics; D041121:Klebsiella oxytoca / Q000201:enzymology* / Q000235:genetics*; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D016133:Polymerase Chain Reaction; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity
D001426:Bacterial Proteins; D017931:DNA Primers; C460086:4-hydroxycinnamate decarboxylase; D002262:Carboxy-Lyases