SOLUBLE AND CELL WALL-ASSOCIATED BETA-GALACTOSIDASES FROM COLD-GROWN WINTER RAPE (BRASSICA-NAPUS L, VAR OLEIFERA L) LEAVES

Soluble and cell wall associated (solubilized from cell wall preparations by 0.5M NaCl) beta-galactosidases were found in non-acclimated or cold-acclimated leaves of winter rape (Brassica napus L., var. L. oleifera L.). Gel filtration of soluble proteins on a Sephadex G-200 column revealed one fraction with beta-galactosidase activity with a molecular weight close to 65 kDa. The temperature optimum for soluble and cell wall-associated beta-galactosidases was 50 degrees C and 55 degrees C, respectively. The cell wall associated form showed higher thermostability at 60 degrees C. Both fractions had a similar pH optimum of 4.0. The K-m against p-nitrophenyl-beta-D-galactopyranoside was 0.25mM and 50mM and the V-max values 2.80 and 1.42 mu mol (mg protein)(-1) min(-1) for the cell wall-associated and soluble form, respectively. The 3-week growth of plants in the cold (2 degrees C) brought about a 22% and 38% decrease in the total and specific activities of the soluble form, respectively. The activity of the cell wall-associated form decreased by ca. 70% during the treatment, independently of the calculation basis. The cold-induced decrease in the enzyme activities was associated with the increase of the K-m value for the soluble form and with the decrease of V-max values for both soluble and wall-associated enzymes. The transient frost(-5 degrees C for 18h followed by 6h at 2 degrees C), applied in darkness, markedly stimulated the activity of the cell wall-associated enzyme in the cold-acclimated leaves. It resulted in decreased K-m and increased V-max values for both soluble and cell wall-associated forms. The possible meaning of cold- and frost-induced changes in the beta-galactosidase activities for cold acclimation of winter rape leaves is discussed.