|
KMID : 0545119980080040353
|
|
Journal of Microbiology and Biotechnology
1998 Volume.8 No. 4 p.353 ~ p.360
|
|
|
Production and Characterization of Acid-stable Pectin Lyase from Bacillus sp. PN33
|
|
Kim, Jong Chon
Kim, Hwa Young/Choi, Yong Jin
|
|
|
Abstract
|
|
|
|
A bacterial strain PN33 producing large amounts of extracellular pectin lyase (PNL, EC 4.2.2.10) was isolated from soil. The isolated bacterium was identified as a strain of Bacillus sp. Production of PNL by the strain was induced only by pectins, with a higher degree of esterification, which had been added to the culture medium as a sole carbon source. The optimal medium for PNL production was determined to consist of 10g pectin, 2g yeast extract, 4 g K_2HPO_4·3H_2O, 0.6 g MgSO_4, and 0.11 g CaCl_2 per liter (pH 7.0). The PNL activity in the culture supernatant reached the highest level of 132 mU/㎖ after 32 h cultivation at 37℃ in the optimal medium. The PNL produced was purified to homogeneity by ammonium sulfate fractionation (50∼80%), and cation exchange and size exclusion chromatographies. The molecular mass of the enzyme was estimated to be approximately 52 kDa by SDS-PAGE. Almost the same mass was determined by nondenaturing PAGE, indicating that the functional enzyme had a monomeric structure. As expected, the PNL exhibited higher activities on the highly esterified pectins whereas it gave no detectable activity on polygalacturonic acid. The enzyme showed the highest activity at the acidic pH of 6.0, exceptional for a bacterial PNL. Maximum activity was measured at 40℃, although the stability of the purified enzyme was poor at this temperature. Calcium (1 mM) was found to activate the PNL activity by 50%, and also remarkably increased the thermal stability of the enzyme. Phenylmethylsulfonylfluoride (PMSF) and diethylpyrocarbonate (DEPC) inhibited the PNL activity almost completely at the concentration of 5 mM. This result indicates that some serine and histidine residues of the enzyme may play an essential role for catalytic function of the enzyme.
|
|
|
KEYWORD
|
|
|
|
|
|
FullTexts / Linksout information
|
|
|
|
|
|
Listed journal information
|
|
  
|
|