Purification and characterization of acid urease from Lactobacillus fermentum.
Acid urease was purified to an electrophoretically homogeneous state, and the molecular weight was estimated to be 220,000. The enzyme consisted of three kinds of subunits, designated alpha, beta and gamma, with molecular weights of 67,000, 16,800 and 8600, respectively, in a (alpha 1 beta 2 gamma 1)2 structure. The isoelectric point of the enzyme was 4.8. The nickel content was found to be 1.9 atoms of nickel per alpha 1 beta 2 gamma 1 unit. The amino acid profile was different from those of known bacterial neutral ureases. The enzyme was most active at pH 2 and around 65 degrees C. It was stable between pH 3 and 9, and below 50 degrees C. The Km for urea was 2.7 mM at pH 2. The enzyme activity was inhibited by Ag+, Hg2+, Cu2+, p-chloromercuribenzoate and acetohydroxamate. The enzyme was separated into three subunits by reverse phase HPLC. The amino terminal amino acid sequences of the subunits alpha, beta and gamma were Ser-Phe-Asp-Met-, Met-Val-Pro-Gly- and Met-Arg-Leu-Thr-, respectively.
Kakimoto
S
S
Microbiology Research Laboratories, Takeda Chemical Industries, Osaka, Japan.
Sumino
Y
Y
Kawahara
K
K
Yamazaki
E
E
Nakatsui
I
I
D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D007700:Kinetics; D007778:Lactobacillus / Q000032:analysis / Q000201:enzymology*; D008670:Metals / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009532:Nickel / Q000032:analysis; D013379:Substrate Specificity; D013696:Temperature; D014508:Urea / Q000378:metabolism; D014510:Urease / Q000302:isolation & purification* / Q000378:metabolism
D004791:Enzyme Inhibitors; D008670:Metals; D009532:Nickel; D014508:Urea; D014510:Urease