Section 9
Chapter 8,905

Intracellular esterase from Lactococcus lactis subsp. lactis NCDO 763: Purification and characterization

Chich, J.-Francois; Marchesseau, K.; Gripon, J.-Claude

International Dairy Journal 7(2-3): 169-174


ISSN/ISBN: 0958-6946
DOI: 10.1016/s0958-6946(97)00001-0
Accession: 008904847

The presence of esterolytic activities in the intracellular extract of Lactococcus lactis subsp. lactis NCDO 763 was investigated. One major activity hydrolyzing beta-naphthyl butyrate was detected. This activity was purified to homogeneity. The enzyme was a homotrimer; the molecular mass of the monomer was estimated by SDS-PAGE to be 29 kDa; by mass spectrometry, it was 29655 +- 30 Da. The pI of the molecule was 4.5. The enzyme was active on para-nitrophenyl esters from C2 to C12 and on orthonitrophenyl butyrate, with a maximum activity on p-nitrophenyl butyrate. The optimum activity on this substrate was found at pH 8.0 and at 55 degree C; kinetic parameters for this substrate were measured at 55 degree C and were K-M = 0.11 mM and V-max = 8.17 mu-mol min-1. The enzyme was strongly inhibited by Pefabloc, diisopropyl fluoro-phosphate and 3,4-dichloroisocoumarin, demonstrating that it was a serine esterase. The amino-terminus of the enzyme was sequenced. The role of this enzyme in cheese ripening is discussed.

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