Isolation and characterization of a tributyrin esterase from Lactobacillus plantarum 2739.
An intracellular tributyrin esterase from Lactobacillus plantarum 2739 was purified to homogeneity by chromatography on DEAE cellulose, Sephacryl 200, carboxymethylcellulose, and Mono Q. The enzyme E2 was separated on DEAE cellulose from a second esterase, E1, and a minor esterase. Additional minor esterases were separated from E2 during chromatography on Sephacryl. E2 was a monomer with a relative molecular mass of approximately 85 kDa. The enzyme was most active at pH 7 and 35 degrees C and retained about 30% of maximal activity at pH 5 and about 18% at 12 degrees C. E2 was strongly inhibited by 1 mM phenylmethylsulfonyl fluoride, Hg2+, or Ag+ and was moderately stimulated by Ca2+ and Mg2+. E2 was active on beta-naphthyl esters of fatty acids from C2 to C10 with a preference for beta-naphthyl butyrate. Tributyrin and, to a lesser extent, tricaprylin and milk fat were also hydrolyzed. Partially purified E1 was more active on tributyrin than was E2. The sequence of the first 15 N-terminal amino acids of purified E2 was Ser-Asn-Glu-His-Thr-Gln-Glu-Val-Leu-Asn-Gln-Thr-Val-Ala-Asp. The enzyme showed a decimal reduction value at 70 degrees C of 2.5 min. The Michaelis constant of E2 on beta-naphthyl butyrate was 0.36 mM.
Gobbetti
M
M
Department of Food Chemistry, University College Cork, Ireland.
Fox
P F
PF
Stepaniak
L
L
D000595:Amino Acid Sequence; D002848:Chromatography, DEAE-Cellulose; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007778:Lactobacillus / Q000201:enzymology*; D008049:Lipase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D008628:Mercury / Q000494:pharmacology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000737:chemistry; D010664:Phenylmethylsulfonyl Fluoride / Q000494:pharmacology; D012834:Silver / Q000494:pharmacology
D004791:Enzyme Inhibitors; D010446:Peptide Fragments; D012834:Silver; D010664:Phenylmethylsulfonyl Fluoride; D008049:Lipase; D008628:Mercury