Characterization of a lysyl aminopeptidase activity associated with phosphoglucose isomerase of Vibrio vulnificus.
Phosphoglucose isomerase (PGI) is a multifunctional enzyme involved in glycolysis and gluconeogenesis and, in mammalian cells, functions as neuroleukin, autocrine motility factor (AMF), and differentiation and maturation factor (MF). We isolated and characterized PGI with a novel lysyl aminopeptidase (LysAP) activity (PGI-LysAP) from Vibrio vulnificus. Mass spectrometry revealed that PGI-LysAP is a heterodimer consisting of 23.4- and 60.8-kDa subunits. Only the heterodimer displayed LysAP activity. PGI-LysAP has a pI around 6.0 and high specificity toward the synthetic, fluorogenic substrate l-lysyl-7-amino-4-methylcoumarin. LysAP activity is optimal at pH 8.0, is 64% higher at 37 degrees C than at 21 degrees C, does not directly correlate with virulence, and is strongly inhibited by serine protease and metalloprotease inhibitors. PGI-LysAP was also identified in Vibrio parahaemolyticus and V. cholerae, but was absent from non-Vibrio human pathogens. Sequencing of the pgi gene revealed 1653 bp coding for a 550-amino-acid protein. Cloned and expressed PGI formed a homodimer with isomerase activity, but not LysAP activity. The finding of LysAP activity associated with heterodimeric PGI should foster a broad search for putative substrates in an effort to elucidate the role of PGI-LysAP in bacteria and its roles in the pathophysiology of diseases.
Richards
Gary P
GP
Agricultural Research Service, US Department of Agriculture, Delaware State University, James W.W. Baker Center, Dover, DE 19901, USA. grichard@desu.edu
Hammer
Carl H
CH
Garfield
Mark K
MK
Parveen
Salina
S
D000595:Amino Acid Sequence; D000626:Aminopeptidases / Q000378:metabolism*; D001426:Bacterial Proteins; D004791:Enzyme Inhibitors; D005956:Glucose-6-Phosphate Isomerase / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D020836:Protein Structure, Quaternary; D021122:Protein Subunits; D013379:Substrate Specificity; D041261:Vibrio vulnificus / Q000201:enzymology* / Q000472:pathogenicity
D001426:Bacterial Proteins; D004791:Enzyme Inhibitors; D021122:Protein Subunits; D000626:Aminopeptidases; C056860:lysyl aminopeptidase; D005956:Glucose-6-Phosphate Isomerase