Dipeptidyl aminopeptidase IV from Pseudomonas sp. WO24.
Dipeptidyl aminopeptidase IV from Pseudomonas sp. WO24 was purified as two molecular forms of 84 and 82-kDa by SDS-PAGE. Peptide mapping and N-terminal sequence analyses indicated that both proteins might be derived from the same protein, and that the 82-kDa molecule might be a truncated form from the 84-kDa molecule at least at the N-terminus. The DAP IV gene of Pseudomonas sp. WO24 was cloned and expressed in E. coli. The enzyme expressed in E. coli JM109 harboring a hybrid plasmid, pYO-6A, with about a 3-kbp fragment containing the DAP IV gene, was purified with an activity recovery of 24%. The recombinant enzyme also had the same two molecular forms, though the ratio of the two forms (about 1:1) was different from that of the native ones (about 1:4). The native and recombinant enzyme preparations had similar specific activities, suggesting that the 84 and 82-kDa molecules are in an active form and have almost the same specific activity. The molecular mass, the subunit number, the substrate specificity, and the effects of various inhibitors of the native enzyme indicated that this enzyme was a typical DAP IV and had properties similar to those of Flavobacterium meningosepticum rather than others.
Ogasawara
W
W
Department of Bioengineering, Nagaoka University of Technology, Niigata, Japan.
Ogawa
Y
Y
Yano
K
K
Okada
H
H
Morikawa
Y
Y
D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D003470:Culture Media; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / Q000235:genetics / Q000302:isolation & purification*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology; D015698:Genomic Library; D007525:Isoelectric Focusing; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010449:Peptide Mapping; D010957:Plasmids; D011549:Pseudomonas / Q000201:enzymology* / Q000235:genetics
D003470:Culture Media; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases